Deformed wing virus (DWV) is a serious pathogen of the honeybee, Apis mellifera L., vectored by the parasitic mite Varroa destructor. The virus is associated with wing deformity in symptomatic bees, and premature death and reduced colony performance in asymptomatic bees. In present study a novel micro PCR-based detection method, termed as ultra-rapid real-time PCR (UR-RT PCR), was developed for the fast and quantitative detection of DWV in honeybee. A specific detection primer set (DWV-UR-F3/R3) was used for the amplification of an unique 133-bp DNA fragment of DWV with a rapid real -time PCR system, GenSpector® TMC-1000, which proceed the cycling with fast heating and cooling rates and a small reaction volume. We showed that this method is able to detect DWV with DNA conditions, artificial recombinant DNA, pBX-DWV479 as well as with virus-infected honeybee samples. In application to a DWV-infected honey bee, the minimum detection time was 8 min 50 seconds under 30 cycles and 10min 11 seconds including melting temperature analysis. This optimizing detection method is one of the fastest real-time PCR-based diagnostic tools and is available to be applied to use for the detection in the field and of various persistency pathogens.

Deformed wing virus(DWV) is one of infectious disease of honey bee that is caused to wings of immature or mutation and death at last. In this study 4 kinds of polyprotein of DWV are compared and then selected 2 kinds of polyprotein, RNA-dependent RNA polymerase(RdRP) and peptidase C3G, which has relatively higher homology than others. Analysis of both RdRP region and peptidase C3G of DWV deposited in Genbank of NCBI showed 99~100% and 95~97% of homology on genomic level, respectively whereas analysis of CRPV-capsid and RNA helicase showed 86~98% and 89~98% of homology, repectively. According to the result of gene analysis, two kinds of polyprotein are constructed and analyzed the homology, resulting in RdRP and Peptidase -C3G showed about 96% and 97% of homology, respectively. It indicated that both region is able to use for generation of specific antibody for the diagnosis of Deformed Wing Virus (DWV).

Deformed wing virus (DWV) of honeybees (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. Here, we report for the first time the occurrence of DWV-infected bumble bees (Bombus terrestris). For the present study, the detection of DWV virus from the adult bumble bee, death adult bumble bee, mail bumble bee, pupa and larva to the infection cycle was investigated in the same colony. Using specific RT-PCR protocols for the detection of DWV followed by sequencing of the PCR products we could demonstrate that the bumble bees were indeed infected with DWV. The virus was detected from diseased insects, and its partial DWV gene was cloned and sequenced. The partial DWV gene encoding the polyprotein is 711-nt of 235 amino acid residues. The deduced nucleotide sequence of the polyprotein partial gene of DWV showed 96.9%, 96.2%, 96.8%, and 96.5% homology to other structure polyprotein partial gene of DWV, respectively. Phylogenetic analysis further conformed that the deduced nucleotide sequence of the polyprotein partial gene of DWV divided to the outside tree. We describe the first time that presence of Deformed wing virus(DWV) from bumble bee(Bombus terrestris) in korea using RT-PCR.

We report for the first time the occurrence of DWV-infected bumble bees (Bombus ignitus). For the present study, the detection of DWV virus from the female and male bumble bee was investigated in the same colony. The Deformed wing virus (DWV) of honeybee (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. Using specific RT-PCR protocols for the detection of DWV followed by sequencing of the PCR products we could demonstrate that the bumble bees were indeed infected with DWV. The virus was detected from Bombus ignitus, and its partial DWV gene was cloned and sequenced. The partial DWV gene encoding the polyprotein is 711-nt of 235 amino acid residues. The deduced nucleotide sequence of the polyprotein partial gene of DWV showed 96.9%, 96.2%, 96.8%, and 96.5% homology to other structure polyprotein partial gene of DWV from insects, respectively. Phylogenetic analysis further conformed that the deduced nucleotide sequence of the polyprotein partial gene of DWV divided to the outside tree. We describe the first time that presence of Deformed wing virus(DWV) from bumble bee(Bombus terrestris) in korea using RT-PCR.