Winter loss of honeybees (Apis Mellifera): Identification of novel and known honeybee viruses associated with weak colonies using state-of-the-art viral metagenomics approaches.

EPILOBEE was a large EU-sponsored project, which was the first to perform honeybee surveillance to better understand honeybee colony losses and other honeybee diseases across 17 European countries. In Belgium, 1261 beehives were sampled during the autumn of 2012 and 2013. After the subsequent winters (spring 2013 and 2014, respectively) the sites were visited again to register the health state of the different selected beehives and to record winter losses of these particular beehives. Prof. Dirk de Graaf, co-promoter of this project, was in charge of the pathogen analysis and therefore these very well characterized samples will be at our disposal for this study.

The Laboratory of Viral Metagenomics (Prof. Jelle Matthijnssens) recently developed a protocol (named NetoVIR) to purify viral particles from stool or other biological samples and sequence them using NGS sequencing technology. A preliminary study using this protocol to investigate the honeybee virome, revealed a plethora of known and surprisingly also unknown novel viruses. The combination of this unique sample set and this novel method will allow us to investigate in great detail the virome of honeybees in Flanders and their relationship with WL.

In practice, samples from 300 Flemish honeybee hives (150 healthy and 150 diseased hives, collected during autumn of 2012 and 2013) will be subjected to our viral metagenomics protocol to identify known and novel viruses. Partially obtained viral genomes of interest will be completed using (RT-)PCR and sanger sequencing. A set of up to 25 viruses of interest will be selected based on their prevalence and likelihood of being honeybee pathogens, and specific q(RT)-PCR assays will be developed. These assays will be used to individually screen each of the 300 selected hives for the presence and viral load of the selected viruses.

The obtained quantitative data on viral loads together with known data about environmental factors and non-viral pathogens (collected during EPILOBEE) will be used to statistically investigate if individual viruses, groups of viruses, viral loads, or viruses in combination with non-viral pathogens or environmental triggers are associated with WL.

Finally we will try to isolate viruses associated with WL using various insect cell lines, or alternatively, perform cage experiments to inoculate virus into adult honeybees or brood for propagation.

Contactpersoon: Prof. Dr. Jelle Matthijnssens

Actor(en): Prof. Dr. Jelle Matthijnssens, Prof. Dr. Dirk C de Graaf

Locatie: KULeuven, Ugent

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