Document Type
Journal Article
Publication Date
8-2015
Journal
PLoS ONE
Volume
10
Issue
8
Inclusive Pages
e0133888
DOI
10.1371/journal.pone.0133888
Keywords
Genes, Reporter; Influenza A virus--genetics; Luminescent Proteins--genetics; RNA Replicase--genetics; Viral Proteins--genetics; Virology--methods
Abstract
Bioluminescent and fluorescent influenza A viruses offer new opportunities to study influenza virus replication, tropism and pathogenesis. To date, several influenza A reporter viruses have been described. These strategies typically focused on a single reporter gene (either bioluminescent or fluorescent) in a single virus backbone. However, whilst bioluminescence is suited to in vivo imaging, fluorescent viruses are more appropriate for microscopy. Therefore, the idea l reporter virus varies depending on the experiment in question, and it is important that any reporter virus strategy can be adapted accordingly. Herein, a strategy was developed to create five different reporter viruses in a single virus backbone. Specifically, enhanced green fluorescent protein (eGFP), far-red fluorescent protein (fRFP), near-infrared fluorescent protein (iRFP), Gaussia luciferase (gLUC) and firefly luciferase (fLUC) were inserted into the PA gene segment of A/PR/8/34 (H1N1). This study provides a comprehensive characterisation of the effects of different reporter genes on influenza virus replication and reporter activity. In vivoreporter gene expression, in lung tissues, was only detected for eGFP, fRFP and gLUC expressing viruses. In vitro, the eGFP-expressing virus displayed the best reporter stability and could be used for correlative light electron microscopy (CLEM). This strategy was then used to create eGFP-expressing viruses consisting entirely of pandemic H1N1, highly pathogenic avian influenza (HPAI) H5N1 and H7N9. The HPAI H5N1 eGFP-expressing virus infected mice and reporter gene expression was detected, in lung tissues, in vivo. Thus, this study provides new tools and insights for the creation of bioluminescent and fluorescent influenza A reporter viruses.
Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.
APA Citation
Spronken, M.I., Short, K., Herfst, S., Bestebroer, T.M., Vaes, V.P. et al. (2015). Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications. PLoS ONE, 10(8): e0133888. doi:10.1371/journal.pone.0133888
Peer Reviewed
1
Open Access
1
Comments
Reproduced with permission of PLoS ONE.