Cargando…
Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications
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 fluores...
Autores principales: | , , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2015
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4524686/ https://www.ncbi.nlm.nih.gov/pubmed/26241861 http://dx.doi.org/10.1371/journal.pone.0133888 |
_version_ | 1782384230564626432 |
---|---|
author | Spronken, Monique I. Short, Kirsty R. Herfst, Sander Bestebroer, Theo M. Vaes, Vincent P. van der Hoeven, Barbara Koster, Abraham J. Kremers, Gert-Jan Scott, Dana P. Gultyaev, Alexander P. Sorell, Erin M. de Graaf, Miranda Bárcena, Montserrat Rimmelzwaan, Guus F. Fouchier, Ron A. |
author_facet | Spronken, Monique I. Short, Kirsty R. Herfst, Sander Bestebroer, Theo M. Vaes, Vincent P. van der Hoeven, Barbara Koster, Abraham J. Kremers, Gert-Jan Scott, Dana P. Gultyaev, Alexander P. Sorell, Erin M. de Graaf, Miranda Bárcena, Montserrat Rimmelzwaan, Guus F. Fouchier, Ron A. |
author_sort | Spronken, Monique I. |
collection | PubMed |
description | 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 vivo reporter 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. |
format | Online Article Text |
id | pubmed-4524686 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-45246862015-08-06 Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications Spronken, Monique I. Short, Kirsty R. Herfst, Sander Bestebroer, Theo M. Vaes, Vincent P. van der Hoeven, Barbara Koster, Abraham J. Kremers, Gert-Jan Scott, Dana P. Gultyaev, Alexander P. Sorell, Erin M. de Graaf, Miranda Bárcena, Montserrat Rimmelzwaan, Guus F. Fouchier, Ron A. PLoS One Research Article 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 vivo reporter 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. Public Library of Science 2015-08-04 /pmc/articles/PMC4524686/ /pubmed/26241861 http://dx.doi.org/10.1371/journal.pone.0133888 Text en © 2015 Spronken et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Spronken, Monique I. Short, Kirsty R. Herfst, Sander Bestebroer, Theo M. Vaes, Vincent P. van der Hoeven, Barbara Koster, Abraham J. Kremers, Gert-Jan Scott, Dana P. Gultyaev, Alexander P. Sorell, Erin M. de Graaf, Miranda Bárcena, Montserrat Rimmelzwaan, Guus F. Fouchier, Ron A. Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title | Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title_full | Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title_fullStr | Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title_full_unstemmed | Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title_short | Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications |
title_sort | optimisations and challenges involved in the creation of various bioluminescent and fluorescent influenza a virus strains for in vitro and in vivo applications |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4524686/ https://www.ncbi.nlm.nih.gov/pubmed/26241861 http://dx.doi.org/10.1371/journal.pone.0133888 |
work_keys_str_mv | AT spronkenmoniquei optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT shortkirstyr optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT herfstsander optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT bestebroertheom optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT vaesvincentp optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT vanderhoevenbarbara optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT kosterabrahamj optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT kremersgertjan optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT scottdanap optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT gultyaevalexanderp optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT sorellerinm optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT degraafmiranda optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT barcenamontserrat optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT rimmelzwaanguusf optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications AT fouchierrona optimisationsandchallengesinvolvedinthecreationofvariousbioluminescentandfluorescentinfluenzaavirusstrainsforinvitroandinvivoapplications |