Deformed wing virus (DWV) is a viral pathogen of the European honeybee (Apis mellifera), associated with clinical symptoms and colony collapse when transmitted by the ectoparasitic mite Varroa destructor. In the absence of V. destructor, DWV infection does not result in visible symptoms, suggesting that mite-independent transmission results in covert infections. True covert infections are a known infection strategy for insect viruses, resulting in long-term persistence of the virus in the population. They are characterized by the absence of disease symptoms in the presence of the virus and by vertical transmission of the virus. To demonstrate vertical transmission and, hence, true covert infections for DWV, a detailed study was performed on the vertical-transmission routes of DWV. In total, 192 unfertilized eggs originating from eight virgin queens, and the same number of fertilized eggs from the same queens after artificial insemination with DWV-negative (three queens) or DWV-positive (five queens) semen, were analysed individually. The F0 queens and drones and F1 drones and workers were also analysed for viral RNA. By in situ hybridization, viral sequences were detected in the ovary of an F0 queen that had laid DWV-positive unfertilized eggs and was inseminated with DWV-positive semen. In conclusion, vertical transmission of DWV from queens and drones to drone and worker offspring through unfertilized and fertilized eggs, respectively, was demonstrated. Viral sequences in fertilized eggs can originate from the queen, as well as from drones via DWV-positive semen.

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