ABC Assay: Method Development and Application to Quantify the Role of Three DWV Master Variants in Overwinter Colony Losses of European Honey Bees

Kevill, Jessica L.; Highfield, Andrea; Mordecai, Gideon; Martin, Stephen J.; Schroeder, Declan C.

doi:10.3390/v9110314

Cited by 77 publications

(109 citation statements)

References 43 publications

(96 reference statements)

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“…We used absolute quantification with DWV-A and DWV-B plasmid standards to accurately determine viral loads in inocula and injected pupae (Supplementary Figure S2). DWV-A and DWV-B RdRp plasmid standards were prepared from the source material cDNA after strain confirmation (above), using the method adapted from Kevill et al (2017). We analyzed the cDNA by PCR with the Kapa2G Robust PCR Kit (Kapa Biosystems), as per manufacturer's instructions, using DWV strain specific RdRp primers ( Supplementary Table S2).…”

Section: Preparation Of Dwv-a and Dwv-b Qpcr Standardsmentioning

confidence: 99%

“…Not all studies distinguish between DWV genotypes. In those studies that do both DWV-A (Highfield et al, 2009;Martin et al, 2012;Mondet et al, 2014;Kevill et al, 2017Kevill et al, , 2019Barroso-Arévalo et al, 2019b) and DWV-B (Natsopoulou et al, 2017) have been associated with colony deaths in the presence of V. destructor. At the same time, high viral loads of DWV-B have been associated with low levels of colony mortality in the United Kingdom and Spain (Mordecai et al, 2016a;Barroso-Arévalo et al, 2019b;Kevill et al, 2019).…”

Section: Introductionmentioning

confidence: 96%

“…Three DWV genotypes have been described: DWV-A, DWV-B, and DWV-C (Mordecai et al, 2016b); only DWV-A (formally DWV) and DWV-B [formally V. destructor virus 1 or VDV-1 (Ongus et al, 2004)] are currently recognized by the International Committee on Taxonomy of Viruses. For clarity, we have employed the types A and B nomenclature widely adopted in recent publications (Martin et al, 2012;McMahon et al, 2016;Mordecai et al, 2016a,b;Brettell and Martin, 2017;Kevill et al, 2017;Gisder et al, 2018;Brettell et al, 2019;Dubois et al, 2019;Kevill et al, 2019;Remnant et al, 2019;Tehel et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

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Accumulation and Competition Amongst Deformed Wing Virus Genotypes in Naïve Australian Honeybees Provides Insight Into the Increasing Global Prevalence of Genotype B

et al. 2020

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Honeybee colony deaths are often attributed to the ectoparasitic mite Varroa destructor and deformed wing virus (DWV), vectored by the mite. In the presence of V. destructor both main genotypes (DWV-A and DWV-B) have been correlated with colony loss. Studies show that DWV-B is the most prevalent genotype in the United Kingdom and Europe. More recently DWV-B has increased in prevalence in the United States. The increasing prevalence of DWV-B at the expense of DWV-A suggests that competition exists between the genotypes. Competition may be due to disparities in virulence between genotypes, differences in fitness, such as rate of replication, or a combination of factors. In this study we investigated if DWV genotypes differ in their rate of accumulation in Australian honeybees naïve to both V. destructor and DWV, and if viral load was associated with mortality in honeybee pupae. We singly and co-infected pupae with DWV-A, DWV-B, and a recombinant strain isolated from a V. destructor tolerant bee population. We monitored viral accumulation throughout pupation, up to 192 h postinjection. We found significant differences in accumulation, where DWV-A accumulated to significantly lower loads than DWV-B and the DWV-recombinant. We also found evidence of competition, where DWV-B loads were significantly reduced in the presence of DWV-A, but still accumulated to the highest loads overall. In contrast to previous studies, we found significant differences in virulence between pupae injected with DWV-A and DWV-B. The average mortality associated with DWV-B (0.4% ± 0.33 SE) and DWV-recombinant (2.2% ± 0.83 SE) injection were significantly less than observed for DWV-A (11% ± 1.2 SE). Our results suggest that a higher proportion of DWV-B infected pupae will emerge into adults, compared to DWV-A. Overall, our data suggest that low mortality in pupae and the ability of DWV-B to accumulate to higher loads relative to DWV-A even during co-infection may favor vector transmission by V. destructor, and may thus be contributing factors to the increasing prevalence of DWV-B globally.

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Section: Preparation Of Dwv-a and Dwv-b Qpcr Standardsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Accumulation and Competition Amongst Deformed Wing Virus Genotypes in Naïve Australian Honeybees Provides Insight Into the Increasing Global Prevalence of Genotype B

et al. 2020

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“…To validate the read-mapping specificity to reliably discriminate between the master variants of DWV, we used two competitive reverse transcription polymerase chain reaction (RT-PCR) assays to target different portions of the virus genome (n = 35 samples from Study 4). Using the ABC assay 45 , which targets the RNA dependent RNA polymerase (RdRp) region in www.nature.com/scientificreports www.nature.com/scientificreports/ DWV, we found a significant correlation between %DWV and DWV type A (Pearson correlation; DWV-A, r = 0.59, p < 0.001, Fig. 6a).…”

Section: Abc and Cba Assays For Dwv-a Subtype Validationmentioning

confidence: 99%

Meta-analysis of honey bee neurogenomic response links Deformed wing virus type A to precocious behavioral maturation

Traniello

Bukhari

Kevill

et al. 2020

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crop pollination by the western honey bee Apis mellifera is vital to agriculture but threatened by alarmingly high levels of colony mortality, especially in Europe and North America. Colony loss is due, in part, to the high viral loads of Deformed wing virus (DWV), transmitted by the ectoparasitic mite Varroa destructor, especially throughout the overwintering period of a honey bee colony. Covert DWV infection is commonplace and has been causally linked to precocious foraging, which itself has been linked to colony loss. taking advantage of four brain transcriptome studies that unexpectedly revealed evidence of covert DWV-A infection, we set out to explore whether this effect is due to DWV-A mimicking naturally occurring changes in brain gene expression that are associated with behavioral maturation. Consistent with this hypothesis, we found that brain gene expression profiles of DWV-A infected bees resembled those of foragers, even in individuals that were much younger than typical foragers. In addition, brain transcriptional regulatory network analysis revealed a positive association between DWV-A infection and transcription factors previously associated with honey bee foraging behavior. Surprisingly, single-cell RNA-Sequencing implicated glia, not neurons, in this effect; there are relatively few glial cells in the insect brain and they are rarely associated with behavioral plasticity. Covert DWV-A infection also has been linked to impaired learning, which together with precocious foraging can lead to increased occurrence of infected bees from one colony mistakenly entering another colony, especially under crowded modern apiary conditions. These findings provide new insights into the mechanisms by which DWV-A affects honey bee health and colony survival. Western honey bees (Apis mellifera) provide an essential pollination service for modern agriculture, with a value estimated at $15 billion per year in the United States 1,2. Extensive reliance on pollination by honey bees has led to great concerns over the massive losses of managed colonies that have occurred since 2006 3-5. These losses are widely thought to be caused by parasites, pathogens, pesticides and poor nutrition, and the main parasite is the ectoparasitic mite Varroa destructor 6-10. Varroa mites are the primary vectors of Deformed wing virus (DWV) 11,12 , a single-stranded, positive-sense RNA virus (family Iflaviridae) that specifically impacts arthropods 13-16. DWV is globally distributed, driving an economic and ecological crisis in honey bee populations 17. DWV was first discovered in symptomatic deformed winged honey bees from Japan in the early 1980s 18 , and the first full genome was sequenced in 2006 19 , hereafter referred to as type A or DWV-A 20. Martin et al. showed through screening the Hawaiian honey bee population that diverse DWV variants persisted prior to the arrival of Varroa 21 ; however, the establishment of Varroa selected for a single variant, DWV-A. DWV-A is currently the most prevalent variant in the North America 22 , thoug...

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“…Most of the recombinant genomic RNA showed similarities with a viral protein-coding sequence derived from VDV-1 and a non-structural proteincoding sequence derived from DWV. A third variant (DWV type C) has more recently been detected in Devon, England (Mordecai et al 2016b) and high viral titers were quantified in collapsing colonies (Kevill et al 2017). However, the prevalence and impact of this DWV variant in the rest of the world are unknown.…”

Section: Introductionmentioning

confidence: 99%

Outcomes of honeybee pupae inoculated with deformed wing virus genotypes A and B

et al. 2019

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Deformed wing virus and Varroa destructor virus-1 have a high percentage of nucleotide identity and might be considered as closely related viruses: DWV genotype A (DWV-A) and DWV genotype B (DWV-B) respectively. They have been implicated in overwinter colony losses in association with Varroa destructor infestations that vectored both DWV variants. In this study, we performed experimental inoculations of honeybee pupae with viral suspensions prepared from honeybee heads naturally infected by either DWV-A or DWV-B. Two outcomes were observed: the inoculated pupae exhibited either higher rates of bees with deformed wings or higher mortality rates than control bees. For both DWV variants, the viral loads quantified in the head of inoculated bees were significantly greater than those in control bees (p < 0.01). These outcomes were not correlated to the virus genotype (DWV-A or DWV-B) detected in the inocula by RT-quantitative PCR targeting the VP3 coding sequence (RT-qPCR). However, the highest mortality rates found in our study were correlated with an increase in sacbrood virus (SBV) load. Despite the fact that only either DWV-A or DWV-B was expected to be inoculated, we observed increased mortality in honeybees that were infected with both DWV and SBV. Deformed wing virus / Varroa destructor virus-1 / virulence / wing deformity / pupae mortality

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ABC Assay: Method Development and Application to Quantify the Role of Three DWV Master Variants in Overwinter Colony Losses of European Honey Bees

Cited by 77 publications

References 43 publications

Accumulation and Competition Amongst Deformed Wing Virus Genotypes in Naïve Australian Honeybees Provides Insight Into the Increasing Global Prevalence of Genotype B

Accumulation and Competition Amongst Deformed Wing Virus Genotypes in Naïve Australian Honeybees Provides Insight Into the Increasing Global Prevalence of Genotype B

Meta-analysis of honey bee neurogenomic response links Deformed wing virus type A to precocious behavioral maturation

Outcomes of honeybee pupae inoculated with deformed wing virus genotypes A and B

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