A real-time PCR method for quantification of the total and major variant strains of the Deformed wing virus

Emma L. Bradford, Craig R. Christie, Ewan M. Campbell, Alan S. Bowman (Corresponding Author)

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Abstract

European honey bees (Apis mellifera) are critically important to global food production by virtue of their pollination services but are severely threatened by deformed wing virus (DWV) especially in the presence of the external parasite Varroa destructor. DWV exists as many viral strains with the two major variants (DWV-A and DWV-B) varying in virulence. A single plasmid standard was constructed containing three sections for the specific determination of DWV-A (VP2 capsid region), DWV-B (IRES) and a conserved region suitable for total DWV (helicase region). The assays were confirmed as specific and discriminatory with limits of detections of 25, 25 and 50 genome equivalents for DWV-A, DWV-B and total-DWV, respectively. The methods were successfully tested on Apis mellifera and V. destructor samples with varying DWV profiles. The new method determined a more accurate total DWV titre in samples with substantial DWV-B than the method currently described in the COLOSS Beebook. The proposed assays could be utilized for the screening of large quantities of bee material for both a total DWV load overview along with more detailed investigations into DWV-A and DWV-B profiles.
Original languageEnglish
Article numbere0190017
Pages (from-to)1-19
Number of pages19
JournalPloS ONE
Volume12
Issue number12
DOIs
Publication statusPublished - 19 Dec 2017

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Deformed wing virus
Viruses
Real-Time Polymerase Chain Reaction
quantitative polymerase chain reaction
Cercopithecine Herpesvirus 1
Bees
Varroidae
methodology
Varroa destructor
viral load
Pollination
Apis mellifera
Honey
Capsid
Assays
Viral Load
Virulence
Limit of Detection
Parasites
Plasmids

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A real-time PCR method for quantification of the total and major variant strains of the Deformed wing virus. / Bradford, Emma L.; Christie, Craig R.; Campbell, Ewan M.; Bowman, Alan S. (Corresponding Author).

In: PloS ONE, Vol. 12, No. 12, e0190017, 19.12.2017, p. 1-19.

Research output: Contribution to journalArticle

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abstract = "European honey bees (Apis mellifera) are critically important to global food production by virtue of their pollination services but are severely threatened by deformed wing virus (DWV) especially in the presence of the external parasite Varroa destructor. DWV exists as many viral strains with the two major variants (DWV-A and DWV-B) varying in virulence. A single plasmid standard was constructed containing three sections for the specific determination of DWV-A (VP2 capsid region), DWV-B (IRES) and a conserved region suitable for total DWV (helicase region). The assays were confirmed as specific and discriminatory with limits of detections of 25, 25 and 50 genome equivalents for DWV-A, DWV-B and total-DWV, respectively. The methods were successfully tested on Apis mellifera and V. destructor samples with varying DWV profiles. The new method determined a more accurate total DWV titre in samples with substantial DWV-B than the method currently described in the COLOSS Beebook. The proposed assays could be utilized for the screening of large quantities of bee material for both a total DWV load overview along with more detailed investigations into DWV-A and DWV-B profiles.",
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note = "Funding: ELB was supported by a Biotechnology and Biological Sciences Research Council (BBSRC) EASTBIO Doctoral Training Partnership (http://www.bbsrc.ac.uk) [grant number BB/J01446X/1] and an Eastern Association Regional Studentship (EARS) and The Morley Agricultural Foundation awarded to ASB. CRC was supported by a KTN BBSRC CASE studentship (BB/M503526/1) (http://www.bbsrc.ac.uk), part-funded by the Scottish Beekeeping Association (https://www.scottishbeekeepers.org.uk/) and the Animal Health - Disease Prevention, Scottish Government awarded to ASB CRC. This project received funding from the European Union’s Seventh Framework Programme for research, technological development and demonstration under grant agreement no 613960 (SMARTBEES) (http://www.smartbees-fp7.eu/) awarded to ASB. The funders had no role in study design, data collection and analysis decision to publish, or preparation of the manuscript. Acknowledgments The authors wish to thank Mr W. Thrale, Mr Z. Blackmore, Mr J. Quinlan, and Mr J. Palombo for sample collection from the South East of England and Margie Ramsey for Beinn Eighe National Nature Reserve sample collection.",
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