Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus
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 (<i>Apis mellifera</i>). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluoresc...
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| Format: | Article |
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MDPI AG
2020-03-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/12/4/374 |
| _version_ | 1797626489889357824 |
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| author | Eugene V. Ryabov Krisztina Christmon Matthew C. Heerman Francisco Posada-Florez Robert L. Harrison Yanping Chen Jay D. Evans |
| author_facet | Eugene V. Ryabov Krisztina Christmon Matthew C. Heerman Francisco Posada-Florez Robert L. Harrison Yanping Chen Jay D. Evans |
| author_sort | Eugene V. Ryabov |
| collection | DOAJ |
| description | 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 (<i>Apis mellifera</i>). 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 <i>egfp</i>-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with <i>egfp</i> 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 <i>egfp</i>-tagged DWV showed gradual accumulation of genome deletions for <i>egfp,</i> providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection. |
| first_indexed | 2024-03-11T10:11:09Z |
| format | Article |
| id | doaj.art-b4048754a20e4f3e9c711adee4cc3304 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-11T10:11:09Z |
| publishDate | 2020-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-b4048754a20e4f3e9c711adee4cc33042023-11-16T14:32:34ZengMDPI AGViruses1999-49152020-03-0112437410.3390/v12040374Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing VirusEugene V. Ryabov0Krisztina Christmon1Matthew C. Heerman2Francisco Posada-Florez3Robert L. Harrison4Yanping Chen5Jay D. Evans6USDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Invasive Insect Biocontrol and Behavior Laboratory, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAUSDA, Agricultural Research Service, Bee Research Lab, Beltsville, MD 20705, USAWe 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 (<i>Apis mellifera</i>). 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 <i>egfp</i>-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with <i>egfp</i> 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 <i>egfp</i>-tagged DWV showed gradual accumulation of genome deletions for <i>egfp,</i> providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.https://www.mdpi.com/1999-4915/12/4/374honey beedeformed wing virusRNA virus vectorinvertebrate virusvirus evolutionpollination |
| spellingShingle | Eugene V. Ryabov Krisztina Christmon Matthew C. Heerman Francisco Posada-Florez Robert L. Harrison Yanping Chen Jay D. Evans Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus Viruses honey bee deformed wing virus RNA virus vector invertebrate virus virus evolution pollination |
| title | Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus |
| title_full | Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus |
| title_fullStr | Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus |
| title_full_unstemmed | Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus |
| title_short | Development of a Honey Bee RNA Virus Vector Based on the Genome of a Deformed Wing Virus |
| title_sort | development of a honey bee rna virus vector based on the genome of a deformed wing virus |
| topic | honey bee deformed wing virus RNA virus vector invertebrate virus virus evolution pollination |
| url | https://www.mdpi.com/1999-4915/12/4/374 |
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