scholarly journals Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 374 ◽  
Author(s):  
Eugene V. Ryabov ◽  
Krisztina Christmon ◽  
Matthew C. Heerman ◽  
Francisco Posada-Florez ◽  
Robert L. Harrison ◽  
...  
Keyword(s):  
Honey Bee ◽  
Essential Gene ◽  
Rna Virus ◽  
Natural Infection ◽  
Fluorescent Microscopy ◽  
Virus Vector ◽  
Cdna Clones ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Leader Protein

We developed a honey bee RNA-virus vector based on the genome of a picorna-like Deformed wing virus (DWV), the main viral pathogen of the honey bee (Apis mellifera). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluorescent protein (eGFP), flanked by the peptides targeted by viral protease, was inserted into an infectious cDNA clone of DWV in-frame between the leader protein and the virus structural protein VP2 genes. The in vitro RNA transcripts from egfp-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with egfp inserts were produced, as evidenced by cesium chloride density gradient centrifugation. These particles were infectious to honey bee pupae when injected intra-abdominally. Fluorescent microscopy showed GFP expression in the infected cells and Western blot analysis demonstrated accumulation of free eGFP rather than its fusions with DWV leader protein (LP) and/or viral protein (VP) 2. Analysis of the progeny egfp-tagged DWV showed gradual accumulation of genome deletions for egfp, providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.

scholarly journals Development of a Honey Bee RNA Virus Vector Based on the Genome of Deformed Wing Virus

2020 ◽  
Author(s):  
Eugene V. Ryabov ◽  
Krisztina Christmon ◽  
Matthew C. Heerman ◽  
Francisco Posada-Florez ◽  
Robert L. Harrison ◽  
...  
Keyword(s):  
Honey Bee ◽  
Fluorescent Protein ◽  
Essential Gene ◽  
Rna Virus ◽  
Natural Infection ◽  
Fluorescent Microscopy ◽  
Virus Vector ◽  
Cdna Clones ◽  
Viral Pathogen ◽  
Deformed Wing Virus

We developed a honey bee RNA-virus vector based on the genome of a picorna-like Deformed wing virus (DWV), the main viral pathogen of the honey bee (Apis mellifera). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluorescent protein (eGFP), flanked by the peptides targeted by viral protease, was inserted into an infectious cDNA clone of DWV in-frame between the leader protein and the virus structural protein VP2 genes. The in vitro RNA transcripts from egfp-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with egfp inserts were produced, as evidenced by cesium chloride density gradient centrifugation. These particles were infectious to honey bee pupae when injected intra-abdominally. Fluorescent microscopy showed GFP expression in the infected cells and Western blot analysis demonstrated accumulation of free eGFP rather than its fusions with DWV LP and/or VP2 proteins. Analysis of the progeny egfp-tagged DWV showed gradual accumulation of genome deletions for egfp, providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.

scholarly journals Development of a honey bee RNA virus vector based on the genome of Deformed wing virus

2020 ◽  
Author(s):  
Eugene V. Ryabov ◽  
Krisztina Christmon ◽  
Matthew C. Heerman ◽  
Francisco Posada-Florez ◽  
Robert L. Harrison ◽  
...  
Keyword(s):  
Honey Bee ◽  
Fluorescent Protein ◽  
Essential Gene ◽  
Rna Virus ◽  
Natural Infection ◽  
Fluorescent Microscopy ◽  
Virus Vector ◽  
Cdna Clones ◽  
Viral Pathogen ◽  
Deformed Wing Virus

AbstractWe developed a honey bee RNA-virus vector based on the genome of a picorna-like Deformed wing virus (DWV), the main viral pathogen of the honey bee (Apis mellifera). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluorescent protein (eGFP), flanked by the peptides targeted by viral protease, was inserted into an infectious cDNA clone of DWV in-frame between the leader protein and the virus structural protein VP2 genes. The in vitro RNA transcripts from egfp-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with egfp inserts were produced, as evidenced by cesium chloride density gradient centrifugation. These particles were infectious to honey bee pupae when injected intra-abdominally. Fluorescent microscopy showed GFP expression in the infected cells and Western blot analysis demonstrated accumulation of free eGFP rather than its fusions with DWV LP and/or VP2 proteins. Analysis of the progeny egfp-tagged DWV showed gradual accumulation of genome deletions for egfp, providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.

scholarly journals Deformed Wing Virus Infection in Honey Bees (Apis mellifera L.)

2019 ◽  
Vol 56 (4) ◽  
pp. 636-641 ◽  
Author(s):  
Roman V. Koziy ◽  
Sarah C. Wood ◽  
Ivanna V. Kozii ◽  
Claire Janse van Rensburg ◽  
Igor Moshynskyy ◽  
...  
Keyword(s):  
Apis Mellifera ◽  
Honey Bee ◽  
Honey Bees ◽  
Rna Virus ◽  
Mandibular Glands ◽  
Deformed Wing Virus ◽  
Clinically Normal ◽  
Bee Health ◽  
Honey Bee Health ◽  
Honey Bee Workers

Deformed wing virus (DWV) is a single-stranded RNA virus of honey bees ( Apis mellifera L.) transmitted by the parasitic mite Varroa destructor. Although DWV represents a major threat to honey bee health worldwide, the pathological basis of DWV infection is not well documented. The objective of this study was to investigate clinicopathological and histological aspects of natural DWV infection in honey bee workers. Emergence of worker honey bees was observed in 5 colonies that were clinically affected with DWV and the newly emerged bees were collected for histopathology. DWV-affected bees were 2 times slower to emerge and had 30% higher mortality compared to clinically normal bees. Hypopharyngeal glands in bees with DWV were hypoplastic, with fewer intracytoplasmic secretory vesicles; cells affected by apoptosis were observed more frequently. Mandibular glands were hypoplastic and were lined by cuboidal epithelium in severely affected bees compared to tall columnar epithelium in nonaffected bees. The DWV load was on average 1.7 × 106 times higher ( P < .001) in the severely affected workers compared to aged-matched sister honey bee workers that were not affected by deformed wing disease based on gross examination. Thus, DWV infection is associated with prolonged emergence, increased mortality during emergence, and hypoplasia of hypopharyngeal and mandibular glands in newly emerged worker honey bees in addition to previously reported deformed wing abnormalities.

scholarly journals A mutualistic symbiosis between a parasitic mite and a pathogenic virus undermines honey bee immunity and health

2016 ◽  
Vol 113 (12) ◽  
pp. 3203-3208 ◽  
Author(s):  
Gennaro Di Prisco ◽  
Desiderato Annoscia ◽  
Marina Margiotta ◽  
Rosalba Ferrara ◽  
Paola Varricchio ◽  
...  
Keyword(s):  
Honey Bee ◽  
Stress Factors ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Mutualistic Symbiosis ◽  
Bee Colony ◽  
Parasitic Mite ◽  
Honey Bee Colony ◽  
Honey Bee Colony Losses

Honey bee colony losses are triggered by interacting stress factors consistently associated with high loads of parasites and/or pathogens. A wealth of biotic and abiotic stressors are involved in the induction of this complex multifactorial syndrome, with the parasitic mite Varroa destructor and the associated deformed wing virus (DWV) apparently playing key roles. The mechanistic basis underpinning this association and the evolutionary implications remain largely obscure. Here we narrow this research gap by demonstrating that DWV, vectored by the Varroa mite, adversely affects humoral and cellular immune responses by interfering with NF-κB signaling. This immunosuppressive effect of the viral pathogen enhances reproduction of the parasitic mite. Our experimental data uncover an unrecognized mutualistic symbiosis between Varroa and DWV, which perpetuates a loop of reciprocal stimulation with escalating negative effects on honey bee immunity and health. These results largely account for the remarkable importance of this mite–virus interaction in the induction of honey bee colony losses. The discovery of this mutualistic association and the elucidation of the underlying regulatory mechanisms sets the stage for a more insightful analysis of how synergistic stress factors contribute to colony collapse, and for the development of new strategies to alleviate this problem.

scholarly journals Deformed wing virus type A, a major honey bee pathogen, is vectored by the mite Varroa destructor in a non-propagative manner

2019 ◽  
Author(s):  
Francisco Posada-Florez ◽  
Anna K. Childers ◽  
Matthew C. Heerman ◽  
Noble I. Egekwu ◽  
Steven C. Cook ◽  
...  
Keyword(s):  
Honey Bee ◽  
Honey Bees ◽  
Varroa Destructor ◽  
Rna Virus ◽  
Fold Increase ◽  
Type A ◽  
Deformed Wing Virus ◽  
Negative Strand ◽  
Viral Loads ◽  
Insect Pollinator

AbstractHoney bees, the primary managed insect pollinator, suffer considerable losses due to Deformed wing virus (DWV), an RNA virus vectored by the mite Varroa destructor. Mite vectoring has resulted in the emergence of virulent DWV variants. The basis for such changes in DWV is poorly understood. Most importantly, it remains unclear whether replication of DWV occurs in the mite. In this study, we exposed Varroa mites to DWV type A via feeding on artificially infected honey bees. A significant, 357-fold increase in DWV load was observed in these mites after 2 days. However, after 8 additional days of passage on honey bee pupae with low viral loads, the DWV load dropped by 29-fold. This decrease significantly reduced the mites’ ability to transmit DWV to honey bees. Notably, negative-strand DWV RNA, which could indicate viral replication, was detected only in mites collected from pupae with high DWV levels but not in the passaged mites. We also found that Varroa mites contain honey bee mRNAs, consistent with the acquisition of honey bee cells which would additionally contain DWV replication complexes with negative-strand DWV RNA. We propose that transmission of DWV type A by Varroa mites occurs in a non-propagative manner.

scholarly journals Tolerance of Honey Bees to Varroa Mite in the Absence of Deformed Wing Virus

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 575 ◽  
Author(s):  
John M. K. Roberts ◽  
Nelson Simbiken ◽  
Chris Dale ◽  
Joel Armstrong ◽  
Denis L. Anderson
Keyword(s):  
Honey Bee ◽  
Honey Bees ◽  
High Throughput Sequencing ◽  
Solomon Islands ◽  
Host Shift ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Bee Health ◽  
Queen Cell

The global spread of the parasitic mite Varroa destructor has emphasized the significance of viruses as pathogens of honey bee (Apis mellifera) populations. In particular, the association of deformed wing virus (DWV) with V. destructor and its devastating effect on honey bee colonies has led to that virus now becoming one of the most well-studied insect viruses. However, there has been no opportunity to examine the effects of Varroa mites without the influence of DWV. In Papua New Guinea (PNG), the sister species, V. jacobsoni, has emerged through a host-shift to reproduce on the local A. mellifera population. After initial colony losses, beekeepers have maintained colonies without chemicals for more than a decade, suggesting that this bee population has an unknown mite tolerance mechanism. Using high throughput sequencing (HTS) and target PCR detection, we investigated whether the viral landscape of the PNG honey bee population is the underlying factor responsible for mite tolerance. We found A. mellifera and A. cerana from PNG and nearby Solomon Islands were predominantly infected by sacbrood virus (SBV), black queen cell virus (BQCV) and Lake Sinai viruses (LSV), with no evidence for any DWV strains. V. jacobsoni was infected by several viral homologs to recently discovered V. destructor viruses, but Varroa jacobsoni rhabdovirus-1 (ARV-1 homolog) was the only virus detected in both mites and honey bees. We conclude from these findings that A. mellifera in PNG may tolerate V. jacobsoni because the damage from parasitism is significantly reduced without DWV. This study also provides further evidence that DWV does not exist as a covert infection in all honey bee populations, and remaining free of this serious viral pathogen can have important implications for bee health outcomes in the face of Varroa.

scholarly journals RNAseq Analysis Reveals Virus Diversity within Hawaiian Apiary Insect Communities

Viruses ◽  
2019 ◽  
Vol 11 (5) ◽  
pp. 397 ◽  
Author(s):  
Brettell ◽  
Schroeder ◽  
Martin
Keyword(s):  
Honey Bee ◽  
Large Scale ◽  
Pest Species ◽  
Insect Communities ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Virus Diversity ◽  
Bee Health ◽  
Honey Bee Health

Deformed wing virus (DWV) is the most abundant viral pathogen of honey bees and has been associated with large-scale colony losses. DWV and other bee-associated RNA viruses are generalists capable of infecting diverse hosts. Here, we used RNAseq analysis to test the hypothesis that due to the frequency of interactions, a range of apiary pest species would become infected with DWV and/or other honey bee-associated viruses. We confirmed that DWV-A was the most prevalent virus in the apiary, with genetically similar sequences circulating in the apiary pests, suggesting frequent inter-species transmission. In addition, different proportions of the three DWV master variants as indicated by BLAST analysis and genome coverage plots revealed interesting DWV-species groupings. We also observed that new genomic recombinants were formed by the DWV master variants, which are likely adapted to replicate in different host species. Species groupings also applied when considering other viruses, many of which were widespread in the apiaries. In social wasps, samples were grouped further by site, which potentially also influenced viral load. Thus, the apiary invertebrate community has the potential to act as reservoirs of honey bee-associated viruses, highlighting the importance of considering the wider community in the apiary when considering honey bee health.

scholarly journals The Gut Microbiota Can Provide Viral Tolerance in the Honey Bee

2021 ◽  
Vol 9 (4) ◽  
pp. 871
Author(s):  
Christopher Dosch ◽  
Anja Manigk ◽  
Tabea Streicher ◽  
Anja Tehel ◽  
Robert J. Paxton ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Honey Bee ◽  
Honey Bees ◽  
Negative Effects ◽  
Positive Role ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Bee Health ◽  
Honey Bee Health

Adult honey bees host a remarkably consistent gut microbial community that is thought to benefit host health and provide protection against parasites and pathogens. Currently, however, we lack experimental evidence for the causal role of the gut microbiota in protecting the Western honey bees (Apis mellifera) against their viral pathogens. Here we set out to fill this knowledge gap by investigating how the gut microbiota modulates the virulence of a major honey bee viral pathogen, deformed wing virus (DWV). We found that, upon oral virus exposure, honey bee survival was significantly increased in bees with an experimentally established normal gut microbiota compared to control bees with a perturbed (dysbiotic) gut microbiota. Interestingly, viral titers were similar in bees with normal gut microbiota and dysbiotic bees, pointing to higher viral tolerance in bees with normal gut microbiota. Taken together, our results provide evidence for a positive role of the gut microbiota for honey bee fitness upon viral infection. We hypothesize that environmental stressors altering honey bee gut microbiota composition, e.g., antibiotics in beekeeping or pesticides in modern agriculture, could interact synergistically with pathogens, leading to negative effects on honey bee health and the epidemiology and impact of their viruses.

scholarly journals Haemolymph removal by the parasite Varroa destructor can trigger the proliferation of the Deformed Wing Virus in mite infested bees (Apis mellifera), contributing to enhanced pathogen virulence

2018 ◽  
Author(s):  
Desiderato Annoscia ◽  
Sam P. Brown ◽  
Gennaro Di Prisco ◽  
Emanuele De Paoli ◽  
Simone Del Fabbro ◽  
...  
Keyword(s):  
Honey Bee ◽  
Varroa Destructor ◽  
Viral Evolution ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Bee Colony ◽  
Bee Health ◽  
Honey Bee Health ◽  
Mite Feeding

AbstractThe association between the Deformed Wing Virus and the parasitic mite Varroa destructor has been identified as a major cause of worldwide honey bee colony losses. The mite acts as a vector of the viral pathogen and can trigger its replication in infected bees. However, the mechanistic details underlying this tripartite interaction are still poorly defined, and, in particular, the causes of viral proliferation in mite infested bees.Here we develop and test a novel hypothesis - grounded in ecological predator-prey theory - that mite feeding destabilizes viral immune control through the removal of both viral ‘prey’ and immune ‘predators’, triggering uncontrolled viral replication. Consistent with this hypothesis, we show that experimental removal of increasing volumes of haemolymph from individual bees results in increasing viral densities. In contrast, we find no support for alternative proposed mechanisms of viral expansion via mite immune-suppression or within-host viral evolution.Overall, these results provide a new model for the mechanisms driving pathogen-parasite interactions in bees, which ultimately underpin honey bee health decline and colony losses.

scholarly journals Home sick: impacts of migratory beekeeping on honey bee (Apis mellifera) pests, pathogens, and colony size

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5812 ◽  
Author(s):  
Samantha A. Alger ◽  
P. Alexander Burnham ◽  
Zachary S. Lamas ◽  
Alison K. Brody ◽  
Leif L. Richardson
Keyword(s):  
Honey Bee ◽  
Pesticide Exposure ◽  
Floral Resources ◽  
Black Queen Cell Virus ◽  
Manipulative Experiment ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Single Location ◽  
Queen Cell ◽  
Bee Colonies

Honey bees are important pollinators of agricultural crops and the dramatic losses of honey bee colonies have risen to a level of international concern. Potential contributors to such losses include pesticide exposure, lack of floral resources and parasites and pathogens. The damaging effects of all of these may be exacerbated by apicultural practices. To meet the pollination demand of US crops, bees are transported to areas of high pollination demand throughout the year. Compared to stationary colonies, risk of parasitism and infectious disease may be greater for migratory bees than those that remain in a single location, although this has not been experimentally established. Here, we conducted a manipulative experiment to test whether viral pathogen and parasite loads increase as a result of colonies being transported for pollination of a major US crop, California almonds. We also tested if they subsequently transmit those diseases to stationary colonies upon return to their home apiaries. Colonies started with equivalent numbers of bees, however migratory colonies returned with fewer bees compared to stationary colonies and this difference remained one month later. Migratory colonies returned with higher black queen cell virus loads than stationary colonies, but loads were similar between groups one month later. Colonies exposed to migratory bees experienced a greater increase of deformed wing virus prevalence and load compared to the isolated group. The three groups had similar infestations of Varroa mites upon return of the migratory colonies. However, one month later, mite loads in migratory colonies were significantly lower compared to the other groups, possibly because of lower number of host bees. Our study demonstrates that migratory pollination practices has varying health effects for honey bee colonies. Further research is necessary to clarify how migratory pollination practices influence the disease dynamics of honey bee diseases we describe here.

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