Recombinant influenza viruses and constructs and uses thereof
Disclosed are compositions and methods comprising one or more recombinant influenza viruses. Recombinant influenza viruses with mutated polymerases and/or rearranged genomes are disclosed. Constructs comprising different influenza nucleic acid sequences are also provided. Methods of inducing protecting immunity with the recombinant influenza viruses are disclosed. Also disclosed are methods of plasmid-free production of influenza virus comprising amplicons comprising one or more of influenza genes.
1. A nucleic acid construct comprising a rearranged influenza type A viral genome segment 2, wherein the rearranged influenza type A viral genome segment 2 comprises:
a) a first influenza type A viral genome segment, wherein the first influenza type A viral genome segment is influenza type A viral genome segment 2 which comprises a nucleic acid sequence that encodes PB1 and
b) a portion of a second influenza type A viral genome segment, wherein the portion of the second influenza type A viral genome segment comprises the portion of influenza type A viral genome segment 8 comprising a nucleic acid sequence that encodes the influenza type A viral protein, NS2, wherein the NS2 nucleic acid sequence is removed from RNA segment 8 of the genome, wherein the NS2 nucleic acid sequence is downstream of the nucleic acid sequence that encodes PB1,
wherein a cleavage site is present between the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence,
wherein the cleavage site is a 2A-like-cis-acting hydrolase element (CHYSEL) site, and
wherein the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence are co-translatable.
2. The construct of claim 1 , wherein the construct further comprises a second rearranged influenza type A viral genome segment comprising a portion of influenza type A viral genome segment 8 comprising at least a portion of a nucleic acid sequence that encodes NS1 and is operably linked to an exogenous sequence.
3. A method of reducing influenza polymerase activity comprising using the construct of claim 1 to produce the influenza by growing the influenza.
4. The method of claim 3 , wherein the rearranged genome comprises an NS2 or M2 nucleic acid sequence operably linked to a PB1 nucleic acid sequence.
5. The nucleic acid construct of claim 1 , wherein the CHYSEL site is a foot-and-mouth disease virus (FMDV) 2A cleavage site.
6. A recombinant type A influenza virus comprising a rearranged influenza type A viral genome segment 2, wherein the rearranged influenza type A viral genome segment 2 comprises:
a) a first influenza type A viral genome segment, wherein the first influenza type A viral genome segment is influenza type A viral genome segment 2 which comprises a nucleic acid sequence that encodes PB1 and
b) a portion of a second influenza type A viral genome segment, wherein the portion of the second influenza type A viral genome segment comprises the portion of influenza type A viral genome segment 8 comprising a nucleic acid sequence that encodes the influenza type A viral protein, NS2, wherein the NS2 nucleic acid sequence is removed from RNA segment 8 of the genome, wherein the NS2 nucleic acid sequence is downstream of the nucleic acid sequence that encodes PB1,
wherein a cleavage site is present between the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence,
wherein the cleavage site is a 2A-like-cis-acting hydrolase element (CHYSEL) site, and
wherein the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence are co-translatable.
7. The recombinant influenza virus of claim 6 , further comprising a rearranged influenza type A viral genome segment comprising an exogenous sequence operably linked to an NS1 sequence.
8. A method of inducing a protective immune response against influenza comprising administering to a subject an effective amount of a composition comprising the recombinant influenza virus of claim 6 , wherein the subject generates a protective immune response against influenza.
9. The method of claim 8 , wherein the recombinant influenza virus comprises an exogenous sequence.
10. The method of claim 9 , wherein the exogenous sequence comprises additional coding and non coding nucleic acid sequences.
11. A vaccine comprising the recombinant influenza virus of claim 6 and a pharmaceutically acceptable carrier.
12. The recombinant type A influenza virus of claim 6 , wherein the CHYSEL site is a foot-and-mouth disease virus (FMDV) 2A cleavage site.