Location: Bee Research LaboratoryTitle: Development of a honey bee RNA virus vector based on the genome of Deformed wing virus
|RYABOV, EUGENE - Non ARS Employee
|CHRISTMON, KRISZTINA - University Of Maryland
|POSADA-FLOREZ, FRANCISCO - Volunteer
|Harrison, Robert - Bob
|Chen, Yanping - Judy
Submitted to: Viruses
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/26/2020
Publication Date: 3/28/2020
Citation: Ryabov, E., Christmon, K., Heerman, M.C., Posada-Florez, F., Harrison, R.L., Chen, Y., Evans, J.D. 2020. Development of a honey bee RNA virus vector based on the genome of Deformed wing virus. Viruses. https://doi.org/10.3390/v12040374.
Interpretive Summary: Deformed wing virus is the most important honey bee virus worldwide, and is a major factor in honey bee declines. It is essential to understand how this virus interacts with its bee host and how these interactions. Here we describe a virus expression system for DWV using a recently designed infectious cDNA clone of genomic RNA of this virus. We use it to show how this virus moves within honey bee cells and to show that this technique can be used to show how viruses attack bees and how the bee immune system responds. These viruses could possibly also be used to deliver medicines or other defenses to honey bees. The results have importance for researchers of bee viruses and possibly longterm benefits for treating bee disease.
Technical Abstract: We developed a honey bee RNA-virus vector based on the genome of a picorna-like Deformed wing virus (DWV), the main viral pathogen of the honey bee (Apis mellifera). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluorescent protein (eGFP), flanked by the peptides targeted by viral protease, was inserted into an infectious cDNA clone of DWV in-frame between the leader protein and the virus structural protein VP2 genes. The in vitro RNA transcripts from egfp-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with egfp inserts were produced, as evidenced by cesium chloride density gradient centrifugation. These particles were infectious to honey bee pupae when injected intra-abdominally. Fluorescent microscopy showed GFP expression in the infected cells and Western blot analysis demonstrated accumulation of free eGFP rather than its fusions with DWV leader protein (LP) and/or viral protein (VP) 2. Analysis of the progeny egfp-tagged DWV showed gradual accumulation of genome deletions for egfp, providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.