Development of a Reverse Genetics System for Toscana Virus (Lineage A)
Toscana virus (TOSV) is a <i>Phlebovirus</i> in the <i>Phenuiviridae</i> family, order <i>Bunyavirales</i>, found in the countries surrounding the Mediterranean. TOSV is an important cause of seasonal acute meningitis and encephalitis within its range. Here, we de...
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doaj-46aac3e9cbb04100b4200e7e43a609f52020-11-25T02:23:40ZengMDPI AGViruses1999-49152020-04-011241141110.3390/v12040411Development of a Reverse Genetics System for Toscana Virus (Lineage A)Akira J. T. Alexander0Marie-Pierre Confort1Sophie Desloire2James I. Dunlop3Srikeerthana Kuchi4Vattipally B. Sreenu5Daniel Mair6Gavin S. Wilkie7Ana Da Silva Filipe8Benjamin Brennan9Maxime Ratinier10Frédérick Arnaud11Alain Kohl12MRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKIVPC UMR754, INRAE, Univ Lyon, Université Claude Bernard Lyon1, EPHE, PSL Research University, F-69007 Lyon, FranceIVPC UMR754, INRAE, Univ Lyon, Université Claude Bernard Lyon1, EPHE, PSL Research University, F-69007 Lyon, FranceMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKIVPC UMR754, INRAE, Univ Lyon, Université Claude Bernard Lyon1, EPHE, PSL Research University, F-69007 Lyon, FranceIVPC UMR754, INRAE, Univ Lyon, Université Claude Bernard Lyon1, EPHE, PSL Research University, F-69007 Lyon, FranceMRC-University of Glasgow Centre for Virus Research, Glasgow G61 1QH, Scotland, UKToscana virus (TOSV) is a <i>Phlebovirus</i> in the <i>Phenuiviridae</i> family, order <i>Bunyavirales</i>, found in the countries surrounding the Mediterranean. TOSV is an important cause of seasonal acute meningitis and encephalitis within its range. Here, we determined the full sequence of the TOSV strain 1500590, a lineage A virus obtained from an infected patient (Marseille, 2007) and used this in combination with other sequence information to construct functional cDNA plasmids encoding the viral L, M, and S antigenomic sequences under the control of the T7 RNA promoter to recover recombinant viruses. Importantly, resequencing identified two single nucleotide changes to a TOSV reference genome, which, when corrected, restored functionality to the polymerase L and made it possible to recover infectious recombinant TOSV (rTOSV) from cDNA, as well as establish a minigenome system. Using reverse genetics, we produced an NSs-deletant rTOSV and also obtained viruses expressing reporter genes instead of NSs. The availability of such a system assists investigating questions that require genetic manipulation of the viral genome, such as investigations into replication and tropism, and beyond these fundamental aspects, also the development of novel vaccine design strategies.https://www.mdpi.com/1999-4915/12/4/411reverse genetics<i>Phlebovirus</i><i>Bunyavirales</i>Toscana virus |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Akira J. T. Alexander Marie-Pierre Confort Sophie Desloire James I. Dunlop Srikeerthana Kuchi Vattipally B. Sreenu Daniel Mair Gavin S. Wilkie Ana Da Silva Filipe Benjamin Brennan Maxime Ratinier Frédérick Arnaud Alain Kohl |
spellingShingle |
Akira J. T. Alexander Marie-Pierre Confort Sophie Desloire James I. Dunlop Srikeerthana Kuchi Vattipally B. Sreenu Daniel Mair Gavin S. Wilkie Ana Da Silva Filipe Benjamin Brennan Maxime Ratinier Frédérick Arnaud Alain Kohl Development of a Reverse Genetics System for Toscana Virus (Lineage A) Viruses reverse genetics <i>Phlebovirus</i> <i>Bunyavirales</i> Toscana virus |
author_facet |
Akira J. T. Alexander Marie-Pierre Confort Sophie Desloire James I. Dunlop Srikeerthana Kuchi Vattipally B. Sreenu Daniel Mair Gavin S. Wilkie Ana Da Silva Filipe Benjamin Brennan Maxime Ratinier Frédérick Arnaud Alain Kohl |
author_sort |
Akira J. T. Alexander |
title |
Development of a Reverse Genetics System for Toscana Virus (Lineage A) |
title_short |
Development of a Reverse Genetics System for Toscana Virus (Lineage A) |
title_full |
Development of a Reverse Genetics System for Toscana Virus (Lineage A) |
title_fullStr |
Development of a Reverse Genetics System for Toscana Virus (Lineage A) |
title_full_unstemmed |
Development of a Reverse Genetics System for Toscana Virus (Lineage A) |
title_sort |
development of a reverse genetics system for toscana virus (lineage a) |
publisher |
MDPI AG |
series |
Viruses |
issn |
1999-4915 |
publishDate |
2020-04-01 |
description |
Toscana virus (TOSV) is a <i>Phlebovirus</i> in the <i>Phenuiviridae</i> family, order <i>Bunyavirales</i>, found in the countries surrounding the Mediterranean. TOSV is an important cause of seasonal acute meningitis and encephalitis within its range. Here, we determined the full sequence of the TOSV strain 1500590, a lineage A virus obtained from an infected patient (Marseille, 2007) and used this in combination with other sequence information to construct functional cDNA plasmids encoding the viral L, M, and S antigenomic sequences under the control of the T7 RNA promoter to recover recombinant viruses. Importantly, resequencing identified two single nucleotide changes to a TOSV reference genome, which, when corrected, restored functionality to the polymerase L and made it possible to recover infectious recombinant TOSV (rTOSV) from cDNA, as well as establish a minigenome system. Using reverse genetics, we produced an NSs-deletant rTOSV and also obtained viruses expressing reporter genes instead of NSs. The availability of such a system assists investigating questions that require genetic manipulation of the viral genome, such as investigations into replication and tropism, and beyond these fundamental aspects, also the development of novel vaccine design strategies. |
topic |
reverse genetics <i>Phlebovirus</i> <i>Bunyavirales</i> Toscana virus |
url |
https://www.mdpi.com/1999-4915/12/4/411 |
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