Background
Methods
Sample and MA104 cells
Fecal sample preparation
RVA isolation
Plaque purifying
RVA identification and growth curve determination
Nuclear acid extraction and whole-genome sequencing
Phylogenetic and genetic analyses
Deduced amino acid sequence alignments and structure-based predictions
Results
Isolation of RVA N4006
Whole-genome sequencing
Phylogenetic and genetic analyses
strain | N1 | N2 | N3 | N4 | N5 | V1 | V2 | V3 | V4 | V6 | V7 |
---|---|---|---|---|---|---|---|---|---|---|---|
CHN/E6398/2019/G9P8-E2 | 98.1 |
99.9
|
100
| 99.9 |
100
|
99.9
|
99.9
|
99.8
| 99.7 |
97.4
|
99.9
|
CHN/Fuzhou18-97/2018/G9P8-E2 | 98.1 | 99.9 |
100
| 99.9 | 99.7 | 99.8 | 99.7 | 99.6 | 99.7 | 97.4 | 99.5 |
CHN/Z2768/2019/G9P8-E2 |
98.2
| 99.8 | 99.9 |
100
|
100
| 99.9 | 99.9 | 99.8 |
99.8
| 97.4 | 99.9 |
CHN/L2448/2019/G9P8-E2 | 97.9 | 99.9 | 99.8 | 99.9 |
100
| 99.8 | 99.6 | 99.7 | 99.7 | 97.4 | 99.7 |
CHN/SZ18442015/2018/G9P8-E2 | 97.9 | 99.9 |
100
| 99.7 |
100
| 99.9 | 99.7 | 99.7 | 99.7 | 97.3 | 99.8 |
CHN/SZ18442205/2018/G9P8-E2 | 98.1 | 99.8 |
100
| 99.9 |
100
| 99.9 | 99.5 | 99.6 | 99.5 | 96.8 | 99.6 |
CHN/Fuzhou19-58/2019/G9P8-E1 | 97.8 |
99.6
| 99.7 | 83.6 | 99.3 |
99.5
|
98.4
|
99.4
|
99.7
|
97.8
|
99.4
|
CHN/Fuzhou19-84/2019/G9P8-E1 |
98.1
| 98.9 | 99.2 |
83.8
| 99.3 | 98.4 | 98.4 | 98.3 | 97.6 | 96.8 | 92.4 |
CHN/LL09131481-2009/G9P8/E1 | 97.2 | 99 |
99.9
| 82.9 | 97.5 | 10.6 | 21.7 | 22.7 | 36.1 | 99.2 | 93.2 |
JPN/Tokyo18-27/2018/G9P8-E2 | 96.8 | 99.8 | 99.9 | 99.9 |
100
| 99.8 |
99.8
| 99.6 | 99.6 |
97.4
| 99.7 |
JPN/Tokyo18-38/2018/G9P8-E2 |
98.2
| 99.7 | 99.9 |
100
|
100
|
99.9
| 99.7 |
99.7
|
99.7
| 97.3 | 99.7 |
JPN/Tokyo18-30/2018/G9P8-E2 | 97.9 | 99.7 | 99.7 | 99.7 |
100
| 99.8 | 99.7 | 99.7 | 99.7 | 97.4 |
99.8
|
JPN/Tokyo18-43/2018/G9P8-E2 | 97.7 |
99.9
|
100
| 99.9 |
100
| 99.9 | 99.7 | 99.7 | 99.7 | 97.4 | 99.8 |
JPN/Tokyo18-50/2018/G9P8-E1 | 97.1 | 99.8 | 99.7 |
83.9
|
100
|
99.8
|
99.8
| 98.2 | 99.7 | 97.4 |
99.7
|
JPN/Tokyo18-37/2018/G9P8-E1 | 97.9 | 98.9 |
99.8
| 83.9 | 99 | 98 | 98.1 | 98.2 | 99.3 |
98.7
| 92.8 |
JPN/Tokyo16-4754/2017/G9P8-E1 | 97.9 |
99.9
| 99.1 | 83.6 | 99.3 | 99.6 | 99.7 |
99.4
|
99.6
| 97.7 | 99.5 |
JPN/Tokyo16-3571/2017/G9P8-E1 | 98.1 | 99.8 | 97.7 | 83.6 | 99.3 | 99.7 | 99.7 | 99.4 | 99.5 | 97.7 | 99.6 |
JPN/Tokyo17-15/2017/G2P4 | 74.4 | 83.3 | 78.3 | 99.9 | 83.8 | 79.6 | 79.3 | 77.6 | 86.4 | 79.6 | 74.6 |
JPN/Tokyo17-17/2017/G2P4 | 75.6 | 83 | 78.3 | 99.6 | 83.7 | 79.8 | 79.7 | 77.1 | 86.3 | 79.5 | 74.5 |
JPN/YR116/2013/G2P4 | 75.8 | 83.2 | 78.4 |
100
| 81.3 | 79.8 | 79.6 | 77.1 | 86.6 | 79.4 | 74.6 |
JPN/KN161/2014/G2P4 | 75.6 | 83.1 | 78.4 |
100
| 83.7 | 79.7 | 79.6 | 77.2 | 86.6 | 79.4 | 74.4 |
Sequence alignments and structure-based predictions
Presumed VP7 antigenic epitope
Strains | 7-1a | 7-1b | 7 − 2 | ||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
87
|
91
|
94
|
96
|
97
|
98
|
99
|
100
|
104
| 123 | 125 | 129 | 130 | 291 |
201
|
211
| 212 |
213
|
238
| 242 | 143 |
145
| 146 |
147
|
148
|
190
|
217
| 221 |
264
| |
N4006 | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
E6398/2019/CHN/G9-E2 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Fuzhou19-58/2019/CHN/G9-E1 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
JZ1810/2018/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Km15119/2016/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D |
T
| I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Km15100/2015/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D |
T
| I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
SC8/2014/CHN/G9 a | T | T | G |
A
| E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
SC6/2013/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
BJ-Q794/2012/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
BJ-Q532/2011/CHN/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Tokyo18-43/2018/JPN/G9-E2 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
NS17-A959/2017/RUS/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
3,000,053,718/2015/USA/G9 a | T | T | G | T | E | W | K | N | Q | D |
T
| I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
3,000,014,466/2014/USA/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
1CR24/2015/THA/G9 a | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Fuzhou19-84/2019/CHN/G9-E1 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E |
G
| G |
Fuzhou18-152/2018/CHN/G9-E2 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E |
G
| G |
BJ-Q33/2010/CHN//G9 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E |
G
| G |
LL09131481/2009/CHN/G9P8-E1 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
L169/2000–2006/CHN/G9 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
CAU10-55/2010/KOR/G9 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E |
G
| G |
SP014/2013/JPN/G9 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
3468/2016/KEN/G9 b | T | T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
3,000,558,285/2016/USA/G9P8 b |
A
| T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
ITA/JES11/2010/G9P8 b | T | T | G | T | E | W | K | N | Q | D | A | I | D | K | Q | N | T | A | D | N | K | D | S | T | L | S | E | S | G |
Rotasiil-AU32/G9 c |
A
| T | G | T | E | W | K |
D
| Q | D | A | I | D | K | Q | N | T | A | D |
T
| K | D | S | T | L | S | E | S | G |
Rotavac-116E/G9 c |
I
| T | G | T | E | W | K |
G
| Q | D | A | I | D | K | Q | N | T | A | D | N | K |
N
| S | T | L | S | E |
N
| G |
RV1/G1 d | T | T |
N
|
G
| E | W | K |
D
| Q | S |
V
|
V
| D | K | Q | N |
V
|
D
|
N
|
T
| K | D |
Q
|
N
| L | S |
M
|
N
| G |
Rotavin-M1/G1 d | T | T |
S
|
G
| E | W | K |
D
| Q |
N
|
V
|
V
| D |
R
| Q | N |
V
|
D
|
N
|
T
| K | D |
Q
|
N
| L | S |
T
|
N
| G |
RV5/G1 d | T | T |
N
|
G
|
D
| W | K |
D
| Q | S |
V
|
V
| D | K | Q | N |
V
|
D
|
N
|
T
| K | D |
Q
|
S
| L | S |
M
|
N
| G |
RV5/G2 d |
A
|
N
|
S
|
D
| E | W | E | N | Q | D |
T
|
M
|
N
| K | Q |
D
|
V
|
S
|
N
|
S
| R | D |
N
| T |
S
|
D
|
I
| S | G |
RV5/G3 d | T | T |
N
|
N
|
S
| W | K |
D
| Q | D | A |
V
| D | K | Q |
D
|
A
|
N
|
K
|
D
| K | D |
A
| T | L | S | E |
A
| G |
RV5/G4 d |
S
| T |
S
| T | E | W | K |
D
| Q |
N
|
L
| I | D | K | Q |
D
| T | A | D |
T
|
R
|
A
| S |
G
|
E
| S |
T
| S | G |
LLR/G10 d | T | T |
N
|
N
| E | W |
T
|
S
| Q |
N
| A |
V
| D | K | Q | N | T |
G
| D |
T
|
R
|
N
| S |
S
| L | S | E |
A
| G |
Presumed VP4 antigenic epitopes
Strains | 8 − 1 | 8 − 2 | 8 − 3 | 8 − 4 | 5 − 1 | 5 − 2 | 5 − 3 | 5 − 4 | 5–5 | ||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
100
|
146
|
148
|
150
|
188
|
190
| 192 | 193 |
194
| 195 | 196 |
180
|
183
| 113 |
114
| 115 |
116
| 125 | 131 |
132
|
133
|
135
|
87
|
88
|
89
|
384
|
386
|
388
|
393
|
394
|
398
|
440
|
441
|
434
|
459
|
429
|
306
| |
N4006 | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Z6398/2019/CHN/G9P[8]-E2 a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Fuzhou18-152/2018/CHN/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
JS2016/2016/CHN/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Km15100/2015/CHN/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
SC9/2014/CHN/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Tokyo18-43/2018/JPN/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
VU12-13-101/2013/USA/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
JES11/2010/ITA/G9P[8] a | D | S | Q | D | S | T | N | L | N | G | I | T | A |
N
| P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
CAU09-376/2017/KOR/G9P[8] a | D | S | Q | D | S | T | N | L | N |
D
| I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Moscow-40/2020/RUS/G3P[8] b | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
Dhaka16/BGW/2003/G1P[8] b | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
CAU17L-103/2009/KOR/G8P[8] b | D | S | Q | D | S | T | N | L | N | G | I | T | A | D | P | V | D | N | R | N | D |
N
| N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
RV1/G1P[8] c | D | S | Q |
E
| S | T | N | L | N |
N
| I | T | A |
N
| P | V | D |
S
|
S
| N | D |
N
| N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
RV5/P[8] c | D | S | Q |
E
| S | T | N | L | N |
D
| I | T | A |
N
| P | V | D | N | R | N | D | D | N | T | N | Y | F |
L
| W | P | G | R | T | P | E | L | R |
Rotavin-M1/P[8] c | D | S | Q |
E
| S | T | N | L | N |
D
| I | T | A |
S
| P | V | D | N | R | N | D | D | N | T | N | Y | F | I | W | P | G | R | T | P | E | L | R |
LLR/P[15] d | D |
T
|
N
|
T
|
Y
| T | N |
Y
|
D
|
S
|
V
| T | A |
P
|
E
|
T
|
T
|
T
|
A
| N |
P
|
Q
|
T
|
S
|
E
| Y | F |
L
| W | P | G | R | T | P |
D
| L | R |
Rotavac-116E/P[11] d |
T
| S |
A
|
A
|
G
|
Y
| N |
V
|
P
|
N
|
A
|
D
|
G
|
A
|
Q
|
T
|
S
|
T
|
D
| N |
S
|
S
|
S
|
N
|
D
| Y | F |
L
| W | P | G |
G
| T | P |
Q
|
C
| R |
RV5/P[5] d |
G
|
A
|
D
| D |
Y
|
V
| N |
Y
|
A
|
S
|
V
| T | A |
T
|
S
|
E
|
T
|
S
|
S
| N |
A
| D |
T
|
G
|
P
| Y | F |
L
| W | P | G | R | T | P | E | L | R |
Discussion
Strain | VP7 | VP4 | VP6 | VP1 | VP2 | VP3 | NSP1 | NSP2 | NSP3 | NSP4 | NSP5 |
---|---|---|---|---|---|---|---|---|---|---|---|
N4006 | G9 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 |
E2
| H1 |
Wa-like RVA | G1 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 | E1 | H1 |
G3 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 | E1 | H1 | |
G4 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 | E1 | H1 | |
G9 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 | E1 | H1 | |
DS-1-like RVA | G2 | P[4] | I2 | R2 | C2 | M2 | A2 | N2 | T2 | E2 | H2 |
Au-1-like RVA | G3 | P[9] | I3 | R3 | C3 | M3 | A3 | N3 | T3 | E3 | H3 |
RVA G9P[8] before 2018 in China | G9 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 |
E1
| H1 |
RVA G9P[8] after 2018 in China | G9 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 |
E1
| H1 |
G9 | P[8] | I1 | R1 | C1 | M1 | A1 | N1 | T1 |
E2
| H1 |