Methods and compositions for increasing replication capacity of an influenza virus
In certain aspects, the present invention provides methods for increasing the replication capacity of influenza viruses in hens' eggs and/or cell culture, recombinant and/or reassortant influenza viruses with increased replication capacity, and immunogenic and vaccine compositions comprising such recombinant and/or reassortant influenza viruses. In other aspects, the invention further provides nucleic acids encoding influenza genes associated with increased replication capacity, expression vectors comprising the nucleic acids of the invention, methods for making influenza viruses with increased replication capacity, and kits useful for practice of the methods.
1. A reassortant influenza virus comprising an M1 protein, wherein the M1 protein comprises an alteration in at least one amino acid, wherein the alteration is at the position corresponding to position 218 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1, and wherein the alteration at the position corresponding to position 218 is to isoleucine.
2. An immunogenic composition comprising the reassortant influenza virus of claim 1 .
3. An influenza vaccine comprising the reassortant influenza virus of claim 1 .
4. The reassortant influenza; irus of claim 1 , wherein the M1 protein further comprises a lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
5. The reassortant influenza virus of claim 1 , wherein the M1 protein further comprises an alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
6. The reassortant influenza virus of claim 1 , wherein the M1 protein further comprises a phenyalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
7. The reassortant influenza virus of claim 1 , wherein the M1 protein further comprises two amino acid alterations selected from the group consisting of:
a. an amino acid alteration to lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and an amino acid alteration to alanine at the position corresponding to position 143 of die M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1;
b. an amino acid alteration to lysine at the position corresponding to position 95 of die M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and an amino acid alteration to phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as show in SEQ ID NO:1; and
c. an amino acid alteration to alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and an amino acid alteration to phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
8. The reassortant influenza virus of claim 1 , wherein the M1 protein further comprises
an amino acid alteration to lysine a the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1;
an amino acid alteration to alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1; and
an amino acid alteration to phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
9. An immunogenic composition comprising the reassortant influenza virus of claim 8 .
10. An influenza vaccine comprising the reassortant influenza virus of claim 8 .
11. A reassortant influenza virus comprising an M1 protein, wherein the M1 protein comprises an isoleucine at the position corresponding to position 218 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
12. An immunogenic composition comprising the reassortant influenza virus of claim 11 .
13. An influenza vaccine comprising the reassortant influenza virus of claim 11 .
14. The reassortant influenza virus of claim 11 , wherein the M1 protein further comprises a lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
15. The reassortant influenza virus of claim 11 , wherein the M1 protein further comprises an alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
16. The reassortant influenza virus of claim 11 , wherein the M1 protein further comprises a phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
17. The reassortant influenza virus of claim 11 , wherein the M1 protein further comprises two amino acid residues selected from the group consisting of:
a. a lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and an alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1;
b. a lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and a phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1; and
c. an alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1 and a phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
18. The reassortant influenza virus of claim 11 , wherein the M1 protein further comprises
a lysine at the position corresponding to position 95 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1;
an alanine at the position corresponding to position 143 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1; and
a phenylalanine at the position corresponding to position 144 of the M1 protein of influenza strain MDV-A as shown in SEQ ID NO:1.
19. An immunogenic composition comprising the reassortant influenza virus of claim 18 .
20. An influenza vaccine comprising the reassortant influenza virus of claim 18 .