3.2 RBD sequence analysis
In view of the role of RBD in the attachment of the virus to ACE2 receptor, neutralization of the virus by antibodies and as a major target for vaccine development, monitoring of RBD sequences over time was undertaken. Table 1 describes mutational analysis of the RBD region over a period of one year (May 2020 to Jun 2021). Till December 2020, none of the samples exhibited characteristic mutations (K417N/T, E484K, N501Y, T478K) documented for the variants of concern identified so far. 2/15 samples from December 2020 did show N440K mutation. In the beginning of March 2021, when the number of cases started showing an upward trend (Fig. 1B), we did not find N501Y mutation characteristic of the UK variant (shared by Brazil and SA variants) as well as K417N mutation in the Brazil/SA variants.
Strikingly, 70% of the NPS specimens sequenced during the beginning of March 2021 and 83% NPS samples sequenced at the end of March exhibited L452R mutation observed in the California variant. In addition, contrary to E484K mutation seen in the strains from Brazil and South Africa, Indian strains exhibited E484Q mutation, defined later as kappa variant. Overall, in the month of March, 33/45 (73%) samples harbour India-specific, L452R/E484Q mutations. Week-wise analysis revealed the same trend, with 70% and 76% of kappa-related cases in 1st and last week of Mar 2021. Simultaneous exponential rise in the number of COVID-19 cases (Fig. 1B) revealed a clear association of emergence of this mutant with the current second wave of the disease. Unfortunately, we did not collect samples during January or February and hence the possibility of earlier detection was missed. As specified in the Table 1, we did identify other unique mutations in a small proportion of NPS specimens.
Analysis of 91 RBD sequences from the samples collected during the month of April led to striking observations. At this time, 34 variants were L452R/E484Q (37%) while 54 sequences harboured L452R/T478K mutations (59%), characteristic of the delta variant. One sequence each exhibited N501Y mutation characteristics of UK strain, N440K mutation and no mutation. Overall, the L452R mutation increased from 0/30 till December to 123/136 (90%) by April (p < 0.001). Importantly, as compared to March (35/45), a significant rise in L452R mutants was seen in April (88/91, p = 0.001). Interestingly at the same time, T478K mutation increased drastically from 4% in March to 59% in April (p < 0.001). The increasing trend of the dominant mutation, L452R/T478K continued in the month of May and June however, we observed significant decline in the number of patients seeking COVID-19 diagnosis at the hospital (Table 1). Our results indicated that the dominant clade G virus strains seems to have been replaced by the variants of concern, L452R/T478K in Pune during the second wave of the disease.
3.3 Analyses based on SARS-CoV-2 full genome sequences from Pune, India
During 2020-21, we sequenced 20 full SARS-CoV-2 genomes from Pune (2020: May: 2, September: 2; 2021: March: 10, April: 2, May: 4). The four mutations, C241T, C3037T, C14408T, and A23403G were observed in all the 20 genomes from the clade “G” (named after the Spike D614G mutation). Of the 10 genomes from March 2021 selected for sequencing, four were wild type while 6 were Kappa variant (B.1.617.1) as per RBD analysis.
As shown in Table 2, two sequences each from May and September 2020 belonged to lineage B.1.1.306 and clade 20B. Of the 10 genomes from March 2021, 2 sequences (211295 – MW969753 and 210761 – MZ021503) belonged to the original prevalent lineage B.1.1.306, clade 20B while the other two sequences, 210922 (MW969752) and 210896 (EPI_ISL_1710598) categorized into separate lineages, B.1.1 (clade 20B) and B.1.36.29 (clade 20A) respectively, suggestive of simultaneous low level of circulation of the wild type virus. Remaining six sequences from March 2021 (210871-MZ021506, 210927-MZ021505, 210929-MZ021504, 211290-MW969754, 211294-MW969755 and 211406-MW969756) formed a distinct lineage B.1.617.1, Kappa variant (clade 21B). Two sequences from Apr 2021 (212095, 212098) and 4 sequences from May 2021 (213366, 213522, 213523 and 213570) formed a distinct clade 21A and belonged to B.1.617.2 lineage (Delta variant, VoC).
The complete genome sequence analysis of sample, 210896 (EPI_ISL_1710598) revealed in-frame stop codon in Orf3a protein at 261 amino acid position (26173 nucleotide position). The first strain with this amino acid change was reported in Germany (hCoV-19/Germany/NW-KRO-2355/2020) in March 2020 and recently in England in June 2021 (hCoV-19/England/HSLL-17FBAD3/2021). It has been reported in 35 countries so far (https://www.gisaid.org/epiflu-applications/covsurver-mutations-app/).
Spike protein
Next, we compared spike protein from the Indian variants with other known variants (Table 2). We found two clusters of Indian variants, B.1.617.1 (Kappa) and B.1.617.2 (Delta) and were distinct from the other variants of concern. With the observed D614G mutation, clade G continued to be the only clade circulating so far in Pune. Mutations specific to the recently emergent variants (UK, South Africa, Brazil and California) were not shared by the Indian variants, however, signature mutations of Delta variant were observed in all six sequences obtained in Apr and May months of year 2021.
Two sequences from 2021 (211295 – MW969753 and 210761 – MZ021503) belonging to the original prevalent lineage B.1.1.306 had four characteristic mutations, L18F, A27S, E484K and Q675H in the spike protein that was missing in all the four sequences from 2020 suggesting that the SARS-CoV-2 is continuously evolving. Two genomes, 210922 (MW969752) and 210896 (EPI_ISL_1710598) had unique mutations at V143F, Q677H and at N440K respectively (Table 2). All the six Indian variants of B.1.617.1 lineage exhibited 5 unique mutations at G142D, L452R, E484Q, P681R, and Q1071H. Additionally, E154K mutation was recorded in 5/6 and V382L, D1153Y mutations were recorded in 4/6 Kappa variants. For Delta variant, all the 6 sequences belonged to B.1.617.2 lineage with 6 unique mutations at T19R, G142D, L452R, T478K, P681R, D950N and deletion at F157 and R158 aa positions with substitution of E156G. Notably, P681R mutation was adjacent to the furin cleavage site (682–685). In-vitro experiments conducted with site-directed mutations have shown that this mutation leads to increase in the fusion activity of SARS-CoV-2 spike protein [14]. Importantly, in the UK variant B.1.1.7, P681H was recorded (Table 2). The acquisition of A222V and K417N in Delta variant (AY.2 lineage) was associated with the upsurge in COVID-19 cases in Europe and USA and these amino acid changes were missing in the Delta variants of this study.
Table 2
Comparison of spike protein mutations in the Indian and other variant viruses (reference: Wuhan strain, accession number, NC045512)
Pangolin Lineages
|
Sequence ID*
|
N-terminal domain in S1 subunit
AA position
|
RBD
AA position
|
C-terminal domain in S1 subunit
AA position
|
S2 subunit
AA position
|
18
|
19
|
20
|
26
|
27
|
70
|
95
|
138
|
142
|
143
|
154
|
171
|
190
|
222
|
382
|
417
|
433
|
440
|
452
|
478
|
484
|
501
|
570
|
614
|
647
|
655
|
675
|
677
|
681
|
716
|
950
|
982
|
1027
|
1071
|
1101
|
1118
|
1133
|
1153
|
1176
|
1251
|
B
|
NC-045512, Wuhan
|
L
|
T
|
T
|
P
|
A
|
V
|
T
|
D
|
G
|
V
|
E
|
V
|
R
|
A
|
V
|
K
|
V
|
N
|
L
|
T
|
E
|
N
|
A
|
D
|
A
|
H
|
Q
|
Q
|
P
|
T
|
D
|
S
|
T
|
Q
|
H
|
D
|
V
|
D
|
V
|
G
|
B.1.1.306
|
8003-IRSHA (MT416725)
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
H
|
|
|
|
|
|
|
|
|
|
|
|
|
8004-IRSHA (MT416726)
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
H
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.1.306
|
CD208550 (MW969758)
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
CD208560 (MW969757)
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
I
|
|
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
V
|
B.1.1
|
CD210922 (MW969752)
|
|
|
|
|
|
|
|
|
|
F
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
H
|
|
|
|
|
|
|
|
|
F
|
|
|
|
B.1.36.29
|
CD210896 (EPI_ISL_1710598)
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
K
|
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.1.306
|
CD211295 (MW969753)
|
F
|
|
|
|
S
|
|
|
|
|
|
|
I
|
|
|
|
|
|
|
|
|
K
|
|
|
G
|
|
|
H
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.1.306
|
CD210761 (MZ021503)
|
F
|
|
|
|
S
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
K
|
|
|
G
|
|
|
H
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.617.1
|
CD210871 (MZ021506)
|
|
|
|
|
|
|
I
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
|
Q
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
H
|
|
|
|
|
|
|
B.1.617.1
|
CD210927 (MZ021505)
|
|
|
|
|
|
|
|
|
D
|
|
K
|
|
|
|
L
|
|
|
|
R
|
|
Q
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
H
|
|
|
|
Y
|
|
|
B.1.617.1
|
CD210929 (MZ021504)
|
|
|
|
|
|
|
|
|
D
|
|
K
|
|
|
|
L
|
|
|
|
R
|
|
Q
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
H
|
|
|
|
Y
|
|
|
B.1.617.1
|
CD211290 (MW969754)
|
|
|
|
|
|
|
|
|
D
|
|
K
|
|
|
|
L
|
|
|
|
R
|
|
Q
|
|
|
G
|
S
|
|
|
|
R
|
|
|
|
|
H
|
|
|
|
Y
|
|
|
B.1.617.1
|
CD211406 (MW969756)
|
|
|
|
|
|
|
|
|
D
|
|
K
|
|
|
|
L
|
|
|
|
R
|
|
Q
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
H
|
|
|
|
Y
|
|
|
B.1.617.1
|
CD211294 (MW969755)
|
|
|
|
|
|
|
|
|
D
|
|
K
|
|
|
|
|
|
|
|
R
|
|
Q
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
H
|
D
|
|
|
|
|
|
B.1.617.2
|
CD212098 (MZ574054)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
V
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
CD213366 (MZ574051)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
CD212095 (MZ574052)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
CD213570 (MZ574053)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
CD213522 (MZ574055)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
CD213523 (MZ574056)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2880930 (India/HR)
|
|
R
|
|
|
|
|
|
|
|
|
|
|
|
V
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2272481 (India/DL)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
V
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2036277 (India/WB)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2897823 (India/MH)
|
|
R
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2956019 (Sweden)
|
|
R
|
|
|
|
|
|
|
D
|
|
|
|
|
V
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.617.2
|
EPI_ISL_2958767 (France)
|
|
R
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
AY.2
|
EPI_ISL_2923711 (USA/CA-CDPH)
|
|
R
|
|
|
|
F
|
|
|
|
|
|
|
|
V
|
|
N
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
AY.2
|
EPI_ISL_2545667 (USA/HI-TAMC)
|
|
R
|
|
|
|
F
|
|
|
D
|
|
|
|
|
V
|
|
N
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
AY.2
|
EPI_ISL_2929274 (USA/CA-CDC)
|
|
R
|
|
|
|
F
|
|
|
D
|
|
|
|
|
V
|
|
N
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
AY.2
|
EPI_ISL_2928214 (USA/CA-OC)
|
|
R
|
|
|
|
F
|
|
|
|
|
|
|
|
V
|
|
N
|
|
|
R
|
K
|
|
|
|
G
|
|
|
|
|
R
|
|
N
|
|
|
|
|
|
|
|
|
|
B.1.1.7 (UK)
|
EPI_ISL_601443
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Y
|
D
|
G
|
|
|
|
|
H
|
I
|
|
A
|
|
|
|
H
|
|
|
|
|
B.1.1.7 (L18F)
|
EPI_ISL_720875
|
F
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Y
|
D
|
G
|
|
|
|
|
H
|
I
|
|
A
|
|
|
|
H
|
|
|
|
|
B.1.1.7 (F490S)
|
EPI_ISL_736026
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Y
|
D
|
G
|
|
|
|
|
H
|
I
|
|
A
|
|
|
|
H
|
|
|
|
|
B.1.1.7 (S494P)
|
EPI_ISL_741039
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Y
|
D
|
G
|
|
|
|
|
H
|
I
|
|
A
|
|
|
|
H
|
|
|
|
|
B.1.1.7 (E484K)
|
EPI_ISL_782148
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
K
|
Y
|
D
|
G
|
|
|
|
|
H
|
I
|
|
A
|
|
|
|
H
|
|
|
|
|
B.1.351 (SA)
|
EPI_ISL_1012924
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
N
|
|
|
|
|
K
|
Y
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
P.1 (Brazil)
|
EPI_ISL_792681
|
F
|
|
N
|
S
|
|
|
|
Y
|
|
|
|
|
S
|
|
|
T
|
|
|
|
|
K
|
Y
|
|
G
|
|
Y
|
|
|
|
|
|
|
I
|
|
|
|
|
|
F
|
|
P.1 (Brazil)
|
EPI_ISL_875566
|
F
|
|
N
|
S
|
|
|
|
Y
|
|
|
|
|
S
|
|
|
T
|
|
|
|
|
K
|
Y
|
|
G
|
|
Y
|
|
|
|
|
|
|
I
|
|
|
|
|
|
F
|
|
P.1 (Brazil)
|
EPI_ISL_875567
|
F
|
|
N
|
S
|
|
|
|
Y
|
|
|
|
|
S
|
|
|
T
|
|
|
|
|
K
|
Y
|
|
G
|
|
Y
|
|
|
|
|
|
|
I
|
|
|
|
|
|
F
|
|
P.1 (Brazil)
|
EPI_ISL_875568
|
F
|
|
N
|
S
|
|
|
|
Y
|
|
|
|
|
S
|
|
|
T
|
|
|
|
|
K
|
Y
|
|
G
|
|
Y
|
|
|
|
|
|
|
I
|
|
|
|
|
|
F
|
|
B.1.427 (California, L452R)
|
EPI_ISL_1620465
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
R
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.429
|
EPI_ISL_824555
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
R
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.525 (E484K)
|
EPI_ISL_1615794
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
K
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.2
|
EPI_ISL_824741
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
B.1.1.432
|
EPI_ISL_913915
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
G
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
*Sequence IDs in bold denote sequences from India; Black: wild type and red: variant viruses |
Table 3
Mutational analyses in nucleocapsid protein with reference to Wuhan strain (accession number, NC045512).
Domains of nucleocapsid protein
|
|
|
N-terminal domain
|
SR-rich
|
Linker region
|
|
Amino acid positions
|
2
|
3
|
63
|
80
|
119
|
142
|
194
|
203
|
204
|
236
|
377
|
Pangolin Lineages
|
Sequence ID*
|
|
|
|
|
|
|
|
|
|
|
|
B
|
NC-045512 (Wuhan)
|
S
|
D
|
D
|
P
|
A
|
P
|
S
|
R
|
G
|
S
|
D
|
B.1.1.306
|
8003-IRSHA (MT416725)
|
|
|
|
|
|
|
|
K
|
R
|
|
|
8004-IRSHA (MT416726)
|
|
|
|
|
|
|
|
K
|
R
|
|
|
B.1.1.306
|
208550 (MW969758)
|
|
|
|
|
|
|
|
K
|
R
|
|
|
208560 (MW969757)
|
|
|
|
|
|
|
|
K
|
R
|
|
|
B.1.1
|
CD210922 (MW969752)
|
|
|
|
|
|
|
|
K
|
R
|
|
|
B.1.36.29
|
CD210896 (EPI_ISL_1710598)
|
P
|
|
|
|
|
|
L
|
|
|
|
|
B.1.1.306
|
CD211295 (MW969753)
|
|
Y
|
|
|
|
S
|
|
K
|
R
|
|
|
B.1.1.306
|
CD210761 (MZ021503)
|
|
Y
|
|
|
T
|
|
|
K
|
R
|
|
|
B.1.617.1
|
CD210871 (MZ021506)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.1
|
CD210927 (MZ021505)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.1
|
CD210929 (MZ021504)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.1
|
CD211290 (MW969754)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.1
|
CD211406 (MW969756)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.1
|
CD211294 (MW969755)
|
|
|
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD212098 (MZ574054)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD213366 (MZ574051)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD212095 (MZ574052)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD213570 (MZ574053)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD213522 (MZ574055)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
CD213523 (MZ574056)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2880930 (India/HR)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2272481 (India/DL)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2036277 (India/WB)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2897823 (India/MH)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2956019 (Sweden)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.617.2
|
EPI_ISL_2958767 (France)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
AY.2
|
EPI_ISL_2923711 (USA/CA-CDPH)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
AY.2
|
EPI_ISL_2545667 (USA/HI-TAMC)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
AY.2
|
EPI_ISL_2929274 (USA/CA-CDC)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
AY.2
|
EPI_ISL_2928214 (USA/CA-OC)
|
|
|
G
|
|
|
|
|
M
|
|
|
Y
|
B.1.1.7 (UK)
|
EPI_ISL_601443
|
|
L
|
|
|
|
|
|
K
|
R
|
F
|
|
B.1.1.7 (L18F)
|
EPI_ISL_720875
|
|
L
|
|
|
|
|
|
K
|
R
|
F
|
|
B.1.1.7 (F490S)
|
EPI_ISL_736026
|
|
L
|
|
|
|
|
|
K
|
R
|
F
|
|
B.1.1.7 (S494P)
|
EPI_ISL_741039
|
|
L
|
|
|
|
|
|
K
|
R
|
F
|
|
B.1.1.7 (E484K)
|
EPI_ISL_782148
|
|
L
|
|
|
|
|
|
K
|
R
|
F
|
|
B.1.351 (SA)
|
EPI_ISL_1012924
|
|
|
|
|
|
|
|
|
|
|
|
P.1 (Brazil)
|
EPI_ISL_792681
|
|
|
|
R
|
|
|
|
K
|
R
|
|
|
P.1 (Brazil)
|
EPI_ISL_875566
|
|
|
|
R
|
|
|
|
K
|
R
|
|
|
P.1 (Brazil)
|
EPI_ISL_875567
|
|
|
|
R
|
|
|
|
K
|
R
|
|
|
P.1 (Brazil)
|
EPI_ISL_875568
|
|
|
|
R
|
|
|
|
K
|
R
|
|
|
B.1.427 (California, L452R)
|
EPI_ISL_1620465
|
|
|
|
|
|
|
|
|
|
|
|
B.1.429
|
EPI_ISL_824555
|
|
|
|
|
|
|
|
|
|
|
|
B.1.525 (E484K)
|
EPI_ISL_1615794
|
|
|
|
|
|
|
|
|
|
|
|
B.1.2
|
EPI_ISL_824741
|
|
|
|
|
|
|
|
|
|
|
|
B.1.1.432
|
EPI_ISL_913915
|
|
|
|
|
|
|
|
K
|
R
|
|
|
*Sequence IDs in bold denote sequences from India; Black: wild type and red: variant viruses |
Nucleoprotein
The Indian variants were distinct in this protein as well (Table 3). The Indian variants did not share the D3L, S236F and P80R mutations seen in the UK and Brazil variants respectively. The two Indian sequences (21295, 210761) had tyrosine (Y) instead of Leucine (L) at 3rd amino acid position as observed in the UK strain. The R203M and D377Y mutation in all the twelve Indian variants. At 203, the change is from hydrophilic Arginine to hydrophobic Methionine while at 377 position, Aspartic acid is changed to Tyrosine. The biological significance of these mutations needs to be evaluated. Remaining Indian sequences continued to harbour R203K and G204R mutations that was present since May 2020. One Indian sequence (210896_EPI_ISL_1710598) was unique with no amino acid substitution at 203 and 204 positions. The six Indian variant exhibited unique mutation at D63G amino acid position as observed in Delta variant.
Other genomic regions:
We compared the other genome regions as well. All the six kappa variant-specific mutations included: Nsp3-T749I, Nsp6-T77A (Orf1a), Nsp12-P323L, Nsp13-M429I, Nsp15-K259R (Orf1b), S26L (Orf3a), I33T (Orf6) and V82A (Orf7a). The other proteins (E, M, Orf7b, Orf8 and Orf10) in Kappa variant remained unchanged. All the six Indian delta variant-specific mutations included: G210T (5’UTR) Nsp2-P129L, Nsp3-P822L, Nsp4-A446V, Nsp6-V149A (Orf1a), Nsp12-P323L, Nsp12-G671S, Nsp13-P77L (Orf1b), S26L (Orf3a), I82T (M), V82A, T120I (Orf7a) and deletions at D (119), F (120) positions in Orf8 region. The other proteins (E, Orf6, Orf7b and Orf10) remained unchanged in the delta variant.