Table 1 Amino acid substitutions or simple point mutations without accumulation of amino acid substitutions by using analysis of whole-genome sequencing of sixth-generation of viruses in each virus strain from both supernatants and cells

Anhui/1

HA

1

C96T

NS

C in P0,occurred in P1-6, T>C in P3

In MDCK1 and MDCK2, both MIX

2

G421A

NS

G>A only in p1-5 A, recovered in P6

Only in MDCK2, MIX

3

G427A

A143T/135*

G in P0,A occurred in P1,A>G in P3,G>A in all P4-6 virus

Complete mutation in MDCK1, MIX in MDCK2

4

C1554T

NS

C in P0, T occurred in P1-2,T>C in P3-6

In both MDCK1 and MDCK2; MIX

1

G503A

N168S/171^#

G in P0-1, A occurred in P2-3, A maintained in P4-6

In both MDCK1 and MDCK2;MIX

NA

2

G996A

G331S/335^#

G in P0-1, A occurred in P2-3 and maintained in P4-6

In both MDCK1 and MDCK2;MIX

3

G1236A

G412K/415^#

G in P0-1, A occurred in P2-3 and maintained in P4-6

In both MDCK1 and MDCK2;MIX

1

G1027A

A343T

G in P0-3, A occurred in P4, A maintained in P5-6

In both MDCK1 and MDCK2; MIX

PA

2

G1039A

A347T

G in P0-3, A occurred in P4, A maintained in P5-6

In both MDCK1 and MDCK2; MIX

3

A1046G

E349G

G in P0-3, A occurred in P4, A maintained in P5-6

In both MDCK1 and MDCK2; MIX

4

C135T

NS

C in P0, T occurred in P1, T maintained in P4-6

In both MDCK1 and MDCK2; MIX

5

C720T

NS

C in P0, T occurred in P1, T maintained in P4-6

In both MDCK1 and MDCK2; MIX

6

C1557T

NS

C in P0, T occurred in P1, T maintained in P4-6

In both MDCK1 and MDCK2; MIX

PB1

1

G226A

D76N

G in P0-3, A occurred in P4, A maintained in P5-6

In only MDCK1, Mix

2

T1356A

V451E

T in P0-3, A occurred in P4, A maintained in P5-6

In only MDCK1 Mix

PB2

1

A260G

D87G

A in P0-3, G occurred in P4, G maintained in P5-6

In only MDCK1 Mix

2

C990A

NS

C in P0, A occurred in P1-3, C maintained in P4-6

In both MDCK1 and MDCK2; MIX

NP

1

G1117A

A373T

G in P0-4, A occurred in P5, A maintained in P6

In both MDCK1 and MDCK2; MIX

2

G1071A

NS

G in P0, A occurred in P1, A maintained in P2-6

In both MDCK1 and MDCK2; MIX

M

1

C900A

NS

C in P0-5, A occurred in P6

In only MDCK2 Mix

Shanghai/2

HA

1

C143T

T48I/40*

C in P0-2,T occurred in P3, T maintained in P4-6

Mix

2

C236T

T79I/71*

C in P0-2,T occurred in P3, T maintained in P4-6

Mix

3

G343A

A117T/107*

G in P0-2,A occurred in P3, A maintained in P4-6

Mix

4

C494T

T165I/157*

C in P0-2,T occurred in P3, T maintained in P4-6

Mix

5

G680A

G227E/218*

G in P0-2,A occurred in P3, A maintained in P4-6

Mix

6

G694A

V237I/223*

G in P0, A occurred in P1,A maintained in P2-4, recovered in P5-6

Mix

7

G1091A

A369K/225*

G in P0-2,A occurred in P3, A maintained in P4-6

Mix

8

T618C

NS

T in P0-3:, T>C in P4:, P5:C>T,P6:T>C

Mix

9

G1011A

NS

P0-3:A>G, P4:G>=A,P5-6 converted to P0(G)

Mix

NA

1

C207T

N73I/72

C in P0, T occurred in P1,T maintained in P2-4, recovered in P5-6

Mix

PA

1

A256G

M86V

A in P0, G occurred in P1,G maintained in P2-3, recovered in P5-6

Mix

2

A484G

Y161A

A in P0,G occurred in P1,G maintained in P2-5,G in P6

Complete mutation in P6

PB1

1

C1544T

S515F

C in P0-2,T occurred in P3,G maintained in P4-6

Mix

2

C1786T

P596S

C in P0-2,T occurred in P3,G maintained in P4-6

Mix

3

C1143T

NS

P0-3:C, P4:C>T,P5-6:T>C

4

C1488T

NS

P0-3:C, P4:C>T,P5-6:T>C

5

G2061A

NS

P0-2:G, P3:G>A,P4:A>G,P5-6:G>A

PB2

1

T554G

I185S

T in P0-P3,G occurred in P4,G maintained in P5-6,G in P6

Complete mutation in P6

2

C393T

NS

P0-3:C>T,P4-6 return to P0(C)

3

C717T

NS

P0-3:C>T,P4-6 return to P0(C)

4

G573A

NS

P0-4:G>A,P5-6 return to P0(G)

NP

1

G850A

A284T

G in P0, A occurred in P1,A maintained in P2-4, A in P5-6

Complete mutation in P5-6

2

G1056A

M352I

G in P0, A occurred in P1,A maintained in P2-4, A in P5-6

Complete mutation in P5-6

3

G987A

NS

P0-6:G>A, G=A in P3,P6 weaken

Mix

4

C1020A

NS

P0-4:C>A, P5-6 converted into P0(C)

Mix

NS

1

G301A

D101N

G in P0-P2, A occurred in P3-4, recovered in P5-6

Mix

2

G622A

E208K

G in P0-P2, A occurred in P3-4, recovered in P5-6

Mix

3

G651A

NS

P0-3:G,P4-6:G>A

Mix