Bunyaviruses with segmented glycoprotein precursor genes and methods for generating these viruses
The invention relates to a bunyavirus, in which separated (NSm)Gn and Gc coding regions are functionally present on two different genome segments, preferably a bunyavirus that comprises a total of at least 4 genome segments. The invention further relates to methods for producing said bunyavirus, and to a composition comprising said bunyavirus and a suitable excipient.
1. A bunyavirus of the genus Phlebovirus , in which separated Gn and Gc coding regions, or separated NSmGn and Gc coding regions, are functionally present on two separate genome segments.
2. The bunyavirus of claim 1 , comprising a total of 4 genome segments.
3. The bunyavirus according to claim 1 , comprising
a bunyavirus L genome segment;
a bunyavirus S genome segment or part of a S genome segment comprising at least the N gene and the 3′ and 5′ UTRs; and
a bunyavirus M genome segment from which the Gn or Gc coding region, or the NSmGn or Gc coding region, has been functionally inactivated,
whereby the Gn or Gc coding region, or the NSmGn or Gc coding region, that is functionally inactivated on the M genome segment is functionally present on a second M genome segment.
4. The bunyavirus according to claim 1 , wherein a bunyavirus L genome segment and/or a S genome segment comprises a foreign gene.
5. A bunyavirus according to claim 1 , wherein the Gn and/or the Gc coding region, or the NSmGn and/or Gc coding region, is from a bunyavirus that differs from the bunyavirus from which the L and S genome segments were obtained.
6. A bunyavirus according to claim 1 , further comprising a nucleocapsid (N) coding region from a bunyavirus that differs from the bunyavirus from which the genome segments and Gn and/or Gc coding region, or the NSmGn and/or Gc coding region, were obtained.
7. A bunyavirus according to claim 1 , wherein an NSs coding region on a S genome segment is functionally inactivated.
8. A method for producing a bunyavirus, the method comprising
A) providing a eukaryotic cell with growth medium; and
B) infecting the eukaryotic cell with the bunyavirus according to claim 1 .
9. A composition comprising a bunyavirus according to claim 1 , and a suitable excipient.
10. The composition according to claim 9 , which is an immunogenic composition.
11. The bunyavirus according to claim 1 , for use as a medicament.
12. A vaccine comprising the bunyavirus according to claim 1 .
13. A method for generating a bunyavirus of the genus Phlebovirus , the method comprising:
A) providing a cell with a bunyavirus of the genus Phlebovirus comprising at least one genome segment functionally encoding a RdRp gene and an N gene, and
B) providing the cell with at least two genome segments selected from L, M, and S genome segments in which separated Gn and Gc coding regions, or separated NSmGn and Gc coding regions, are functionally present on two separate genome segments.
14. The method according to claim 13 , whereby the cell is provided with a genomic segment by providing the cell with a vector that comprises cDNA of said genomic segment which is flanked by a T7 promoter and cDNA of a ribozyme, further comprising providing the cell with a T7 polymerase.
15. The bunyavirus of claim 1 , comprising a total of 3 genome segments.
16. The method according to claim 13 , wherein the separated Gn and Gc coding regions are present on two separate minigenome segments, or on a genome segment and a minigenome segment.
17. The composition according to claim 9 , for use as a medicament.
18. A vaccine comprising the composition according to claim 9 .