Update on geographical variation and distribution of SARS-nCoV-2: A systematic review

Abstract

Knowledge of a new mutant strain of SARS-coronavirus (CoV-2) is enormously essential to identify a targeted drug and for the development of the vaccine. In this article, we systematically reviewed the different mutation strains (variant of concern [VOC] and variant of interest [VOI]) which were found in different countries such as the UK, Singapore, China, Germany, Vietnam, Western Africa, Dublin, Ireland, Brazil, Iran, Italy, France, America, and Philippines. We searched four literature databases (PubMed, EMBASE, NATURE, and Willey online library) with suitable keywords and the time filter was November 2019 to June 16, 2021. To understand the worldwide spread of variants of SARS-CoV-2, we included a total of 27 articles of case reports, clinical and observational studies in the systematic review. However, these variants mostly spread because of their ability to increase transmission, virulence, and escape immunity. So, in this paper is we found mutated strains of SARS-CoV-2 like VOCs that are found in different regions across the globe are ALPHA strain in the U.K, BETA strain in South Africa, GAMMA strain in Brazil, Gamma and Beta strains in European Countries, and some VOIs like Theta variant in the Philippines.

Introduction

Coronavirus is a disease of high infectivity and pathogenicity, which has been evolved with time as SARS-coronavirus (CoV), Middle East respiratory syndrome-CoV (MERS-CoV), and SARS-nCoV-2 in 2003, 2012, and 2019, respectively.[1] It is a danger to people's lives and health as the confirmed cases of COVID-19 globally are 175,686,814 of which India has 29,510,410 cases and this virus has caused 3,803,592 deaths worldwide since the start of the deadly disease.[2] The structure of the largest RNA virus (coronavirus) genome has genes in the sequence from 5' to 3' end in the order of ORF1a, ORF1b, S, ORF3, E, M, and N among which 0.6% is ORFa/b genes.[3] Sixteen nonstructural proteins (NSPs) are originated from two polyproteins or viral replicases proteins, PP1a and PP1ab that are produced by ORFa/b genes.[4] The other half of the sequence produces spike proteins, envelope proteins, membrane proteins present at the outer surface, and nucleocapsid protein present with the RNA genome. These all proteins are known as structural proteins.[5] The receptor-binding domain (RBD) and 12 nucleotides inserted at S1/S2 site (furin-like cleavage site, NSPRRAR) domain make the unique structure of SARS-CoV-2 that is unlike other existing corona-viruses.[6]

Recombination occurs with the change in the structure and functions of the proteins that lead to the alteration in the virulence, transmissibility, and probability to escape naturally and vaccine-induced immunity that can further increase the risk of severe infection[5,7] Because of the high recombination rate,[8] several case studies, clinical studies, and observational studies have shown different variants of Coronavirus at the global level carrying mutations in ORF1a, furin cleavage site Nucleocapsid protein, and in M gene. Mutations in spike proteins have shown increased transmissibility and some of the significant mutations seen are D614G mutation. According to the different variations of the strains globally, the virus evolution group of the WHO has founded a new naming format for the variants of SARS-nCoV-2 to ease the recognition and communication of these variants.[9] If the mutations in the genome of SARS-CoV-2 leads to any phenotypic changes and have caused cluster of cases that further leads to community transmission are referred to as variant of interest (VOI) and if this VOI has shown increased transmissibility, virulence or decreased the effect of therapeutics; then it is referred as variant of concern (VOC).[10] With the maximum public health implications, some of the current VOC for SARS-CoV-2 are alpha, beta, gamma, and delta that are detected first in United Kingdom, South Africa, Brazil, and India, respectively.[9,10,11]

In our study, we have also discussed the other Geographical variations in the SARS-CoV-2 genome that are reported in UK, Singapore, China, Germany, Vietnam, Western Africa, Dublin, Ireland, Brazil, Iran, Italy and France, America, Philippines.

Materials and Methods

We screened five literature databases (PubMed, Embase, Nature, Willey online library), using the keywords: “coronavirus disease,” “SARS-CoV-2,” “covid19,” “mutation,” “polymorphism,” “variant,” “mutant,” deletion,” “duplication” and Time filter of 2019 November to June 16, 2021. First, we screened all the abstracts and titles of the articles and after that, we selected relevant articles based on predefined exclusion and inclusion criteria by reading full texts of the articles.

Inclusion-exclusion criteria

We included clinical studies, case reports, and observational published studies which are evaluating the different mutation strains of SARS-CoV-2 globally. Only COVID-19 positive patient studies were considered and included in the clinical studies for analyzing and identifying the different new mutations of SARS-CoV-2. The total number of studies that we researched was 120, out of which 27 were included and 93 were excluded depending on the criteria that we selected. Preclinical studies, COVID-19 associated diseases studies, other studies related to mutations caused in the genome because of any illness or diseases are neglected and defines the exclusion criteria for our study.

Results

The four-literature database was preliminary screened out then we found a total of 120 nonduplicated studies after the screening of title and abstract, a total of 27 [Table 1] articles were selected for further full-text screening which were satisfying the inclusion criteria and 93 articles were excluded (4 preclinical studies, 8 nonmutational COVID-19 studies, 39 COVID-19 associated studies and 42 other studies according to exclusion criteria. The PRISMA flow chart of own presented in Figure 1.

Table 1.

Geographical characterization of mutations found in severe acute respiratory syndrome coronavirus 2 patients

Country	Protein	Mutation	Published
United Kingdom	Spike protein	D614G mutation	Weissman et al.[12], Sekulic et al.[13]
		D796H, ΔH69/ΔV70, Y200H and T240I, P330S, W64G, D796H, ΔH69/ΔV70	Steven A Kemp, 2021[14]
		B.1.1.7 lineage (N501Y), (69-70 deletion and 144del) N501Y, AS70D, D614G, P681H, T716I, S982A and D1118H (A222V and D614G)	Dao et al.[15]
		T739I	Sekulic et al.[13]
		H69R, N74K, N99KLNY, H245R, S247R, T259K, G261R, T307I	Ko et al., 2021[16]
	Furin cleavage site regions	R682L, S686G
	M gene	T7I substitution
	Orf1b	variant 14407 in specimen A and variants 14407 and 14408 in specimen B	Tillett et al.[17]
		P314L	Sekulic et al.[13]
	ORF8	Q27stop, R52I, and Y73C	Dao et al.[15]
	Neucleocaspid protein	D3L, R203K, G204R, and S235FA220V
	NSp3	M125I
	NSP12	P323L
	ORF1a	L3606F, T265I, L3083I, A3958_T3959delAT	Sekulic et al.[13]
	ORF3a	Q57H
Singapore	ORF8	∆382 deletion	Young et al.[18]
	Spike	D614G mutation	Young et al., 2021[19]
Zhejiang China	Furin cleavage site	35 specific gene mutations Ser 596, Gln613, Glu702, Ala771, Ala1015, Pro1053, Thr1066	Jin et al.[20]
Germany	N gene	SNP at the N terminal side (N terminal C28858T, middle C29200T, C term CC29451T)	Ziegler et al.[21]
Vietnam	ORF1ab coding sequence	Nonsynonymous mutations: G8388A (Ser to Asn), A8987T (Ile to Phe) and C10232T (Arg to Cys). A synonymous mutation: G3778A	Phan et al., 2020[22]
Benin, Western Africa	Spike protein	D614G	Sander et al., 2021[23]
South Africa		B.1.351 (K417N, E484K, N501Y)	SARS-CoV-2, 2021[24]
Dublin, Ireland	N gene	GGG to AAC mutations at the position 28,881, 28,882, 28,883, 28,821	Lee et al., 2021[25]
Brazil	Spike protein	K417T, E484K, N501Y. (Pangolin lineage P. 1) P. 1 lineage	Fujino et al., 2021[26]
			Siqueira et al., 2021[27]
	-	5 SNV	Siqueira et al., 2021[28]
	Spike	P. 1 lineage	Romano et al., 2021[29]
Iran	5’UTR	241 C>T nucleotide241 C>T nucleotide	Salehi-Vaziri et al., 2021[30]
	ORF1ab, NSP3	3037 C>T nucleotide
	ORF1ab, NSP12	14408 C>T nucleotide
	ORF1ab, NSP14	18877 C>T nucleotide
	Spike	23403 A>G nucleotide
	NS3	25563 G>T nucleotide
	Nucleocapsid	28854 C>T nucleotide
	M (matrix)	26735 C>T nucleotide
Italy	Orf1ab	N79S, A526L	Monne et al., 2020[31]
	ORF6	C>T at the nt. 27367	Delbue et al., 2021[32]
	Spike	P. 1 (K417T, E484K, and N501Y)	Maggi et al., 2021[33]
		B.1.351	Novazzi et al., 2021[34]
France	Spike	N501Y and E484K (B.1.351)	SARS-CoV-2, 2021[24]
America	Spike	D614 G	Díaz et al.[35]
Philippines	Orf1ab gene	S1188L, K1795Q, S3675-F3677del	Fabiani et al.[36]
	ORF8	E92K
	ORF3a	G174C
	ORF1ab	A1809V, V4225L, P4715L, D5130Y, and E5665D
	Spike	D138Y, R190S, K417T, E484K N501Y, H655Y, T1027I, D614G, S640F, V1176F
	N gene	P80R, RG203KR

SARS-CoV-2 – severe acute respiratory syndrome coronavirus 2

Figure 1.

PRISMA flow chart of the study

Different mutated strains of SARS-nCoV-2 at the global level

After studying the full text of clinical studies, case reports, and observational studies, we could find out the different mutated strains of SARS-CoV-2 that are extended at the global level as summarized in Table 1. Due to very high recombination rate of the virus, different countries (Such as in United Kingdom, Singapore, China, Brazil, Vietnam, Italy, Germany, Africa, Ireland, Iran, France, America, and Philippines) have different variants of SARS CoV 2 that are expressing several mutations found in the patients.

Discussion

In our study, we found that a variety of mutations documented in the coronavirus (SARS-CoV-2) are due to high recombination rate and are found in spike protein, furin cleavage site, nucleocapsid protein, ORF genes (ORF1ab, ORF8, ORF1a, ORF3b, ORF6), NSP3, NSP12, NSP14 and in M gene. Among all the mutations, predominantly the mutations in spike protein are noted in maximum regions of the world and spike proteins help in the entry of coronavirus into the host cell via interaction with the angiotensin-converting enzyme 2 (ACE2) domain.[37]

Spike protein mutations

The spike protein mainly comprises S1 subunit and S2 subunit. The S1 subunit has different four domains that are A, B, C, D domains. A domain refers to the N terminal domain that recognizes carbohydrate which are required for the attachment with the cell receptor of host cells. However, the B domain of spike protein is also known as RBD helps in the interaction with ACE-2 host cell receptor.

The unique site of SARS-CoV-2 that is the FC site (furin cleavage site) is located between S1 and S2 domains and is comprised the PRRA sequence. In the S2 subunit, a cleavage site (S2') and a C terminal is present. Both these cleavage sites that is furin cleavage and S2' cleavage site help the virus to enter into the host cell.[38]

New variant of SARS-CoV-2 carrying a variety of mutations in S protein such as in case of the United-Kingdom mutation in B.1.1.7 lineage (Alpha strain), D614G mutation, and other mutations like D796H, ΔH69/ΔV70, Y200H, T240I, P330S, W64G, D796H, ΔH69/ΔV70, N501Y, A570D, P681H, T716I, S982A, D1118H, A222V, T739I, H69R, N74K, N99KLNY, H245R, S247R, T259K, G261R, and T307I were found. In the alpha strain, spike protein mutations as N501Y substitution and deletion from 69 to 70, results in increased transmission rates of the virus to about 74% more than other variants.[15]

According to our study, D614G mutations in spike proteins are prevailing in Western Africa, America, and Singapore and this mutation is more infectious.[39] In the comparative study, the Glycine 614 was found to be more efficient than aspartic acid 614 in D614G mutations of spike proteins.[12,40] Mutations in P. 1 lineage (K417T, E484K, and N501Y) that is also referred to as Gamma strain of SARS-CoV-2 which are reported in Brazil and Italy,[23,26,27,29] B.1.351 (Beta strain) with mutation on the position of K417N, E484K, and N501Y are found in Africa, Italy,[34] and France.[24] In South Africa, B.1.351 major outbreak variants of novel coronavirus were found.[34]

Hence, there is recently three different types of variants identified Alpha (N501Y), Beta (K417N, E484K, and N501Y) and gamma variant (K417T, E484K, N501Y) in RBD of S protein of Corona virus-2. All three mutant variant were shown more transmissibility.[41] In the Philippines, some of the mutations are reported like R190S, E484K, D138Y, K417T, N501Y, H655Y, T1027I, D614G, S640F, V1176F in the spike region of SARS-CoV-2 that are representing it as to be the theta strain.[36]

All lineage strains of SARS-CoV-2 which was found in our study are presented in tabulated form in Table 1. The study given information of different variants of mutation present in the spike protein of SARS-corona virus-2 which were recognized by sequencing and other methods in patient studies worldwide. Here, we present the major geographical locations of the variants, positions of the mutation sites, the different mutation types observed, presence of multiple mutation in S protein and RBD region which make interaction with host cell ACE2 receptor.[38] This study helps in developing and designing therapeutic target drugs and vaccines to overcome the burden of COVID-19 and its variants.

Furin cleavage binding site mutations

The unique sequence of SARS-CoV-2 consists of polybasic sequence, PRRAR also known as furin cleavage site, present between the S1 and S2 site and is identified by phylogenetic analysis.[42] This spike protein cleaved by furin on the S1/S2 site and that further allows the entry of the virus and mediates the membrane fusion. However, a number of spike protein mutation can modulate the efficacy of furin cleavage.[43] In human clinical samples, the furin cleavage binding site mutations at R682 L, S686G were found in the United Kingdom[16] and in China, Ser596, Glu702, Gln613, Ala771, Ala1015, Pro1053, Thr1066 mutations with other 35 specific gene mutations were recognized in human blood samples.[20] Moreover, ZJ01 mutations were also identified near the Furin cleavage site (FC site) that affect the binding capability of Furin.[20]

Membrane (M) protein mutations

The M protein of corona-virus plays an important role in virus assembly and helps to make a new virus particles.[44] Membrane (M) protein also binds with Nucleocapsid protein for stabilization.[45] According to sung heeko, T7I substitution mutation are found in M gene[16] and another study identified mutation on 26735 C > T nucleotides.[30]

Nucleocapsid protein mutations

N protein (nucleocapsid protein) is a complex of RNA-binding protein which plays an important role in the replication and transcription process of viral particles.[46] According to Manh H. Dao, nucleocapsid proteins was mutated on D3 L, R203K, G204R, S235F and A220V in UK patients.[15] In Germany, SNP were found at each terminal of M-protein.[25] In Ireland, GGG-to-AAC mutation at the position 28881, 28882, 28883, 28821 were found[24] and in Philippines P80R, RG203KR mutations were reported in N region.[36]

ORF region mutations

SARS-CoV-2 contain positive-strand RNA genome which has seven gene sequences in order of the following 5' to 3'direction ORF1a, ORF1b, S, ORF3, E, M, N. the ORF1ab gene are translated into PP1a and pp1ab polypeptide and processed by protease to produce 1–16 NSP protein.[4,47,48]

Missense mutation in ORF1a at the amino acid L3606F, T265I, L3083I and frameshift deletion (A3958_T3959delAT)[13] and also other mutations such as P314 L,[13] R52I, Y73C mutation in ORF8[15] were found in the United Kingdom. In Singapore, Δ382 deletion in ORF8 were found in the COVID-19 patients.[18] In Vietnam, according to Lan T. Phan, 2020, nonsynonymous mutations in coding sequence “G8388A” (Ser to Asn), A8987T (Ile to Phe), and C10232T (Arg to Cys) were reported. However, Synonymous mutations in coding sequence G3778A and N79S, A526 L mutations were reported in Italy,[22] orf1ab mutation (N79S, A526 L)[31] and ORF1ab mutation at nucleotides were also recognized in Iran.[30] In the Philippines, orf1ab gene mutation on S1188 L, K1795Q, S3675-F3677del, ORF8 mutated on E92K, ORF3a mutated on G174C, ORF1ab mutated on A1809V, V4225 L, P4715 L, D5130Y, and E5665D amino acid position.[36]

Nonstructural proteins mutations

In the case of the UK, NSP3 mutated on M125I amino acid and NSP12 mutated on P323 L.[15] In Iran, mutations at NSP3 nucleotides are reported[30] NSP3 helps in the translation process of viral protein and suppress the synthesis of the host cell protein whereas NSP12 helps in RNA polymerase/replicase activity.[47]

This study informs us about the different types of mutations that were identified in spike proteins, ORF gene, N protein, M protein, and nucleocapsid in SARS-CoV-2 which were recognized mainly by sequencing and reverse transcription-polymerase chain reaction in COVID-19 patients of different countries. Here, we present geographical locations, mutation sites in proteins, and genes. We found some mutant strains that are alpha, beta, gamma considered as variants of concerns [Table 2] as these are highly spreading in most of the countries. This study helps in developing therapeutic target drugs and designing vaccines to overcome the burden of COVID-19 and its variants.

Table 2.

This table concludes the variant of concerns that are of the lineage B.1.1.7, B.1.351, P.1 and B.1.617.2 and named as alpha, beta and gamma by WHO. These variants of concerns are found in our studies and the implications caused by these mutations in SARS-CoV-2 are also tabulated

WHO label	Variant type	County of first detection	Pangolineage/GISIAD clade or variant	Mutation implications	References
Alpha	VOC	United Kingdom	B.1.1.7 lineage3 deletions (69-70del and 144del)(N501Y),(N501Y, A570D, D614G, P681H, T716I, S982A, and D1118H), A222V and D614G mutations	Highly transmissible	[14]
Beta	VOC	South Africa	B.1.351 (20H/501Y.V2) N501Y and E484K mutations	Higher transmissibility and immune escape	[23]
			K417N, E484 K, and N501Y	2.7 and a 6.4-fold geometric mean reduction in neutralizationResistance to convalescent plasma and several mAbs	[33]
Gamma	VOC	Brazil	P. 1 (20J/501Y.V3)	Escape naturally and vaccine-induced immunityIncreased viral load with increased risk of severe infection or higher mortality and might have <70% increased transmissibility	[42]
			K417T, E484K, and N501Y	Unknown implications	[32]

These variants of concerns are found in our studies and the implications caused by these mutations in SARS-CoV-2 are also tabulated. SARS-CoV-2 – severe acute respiratory syndrome coronavirus 2; VOC – Variant of concern

Conclusion

This study concludes that alpha, beta, and gamma VOCs that are originated from the United Kingdom, South Africa, and Brazil, respectively, are reported in clinical studies, observational studies, and case reports across the globe. These variants are spreading because of their capability, of high transmissibility, to escape natural and vaccine-induced immunity and to increase the viral load that further enhances the risk of severe infection and higher mortality rate. With these variants of concerns, there is one more VOC that is named as Delta variant (B.1.617.2), originated in India, known to increase the transmissibility of SARS-CoV-2[11,49]

There are other mutations at different regions of the SARS-CoV-2 (spike protein, ORF8 gene, N gene, M gene, Furin cleavage site, etc.) that are reported in our study. VOIs that is theta variant originated in the Philippines is also reported in the study. The significance of this study is to focus on the targets at the different regions of the virus for incorporating the mutated strains of the virus for therapeutic utilization like vaccine development and drug discovery. Moreover, to fight with the virus that has a high recombination rate, booster shots of vaccines may be required with time with different trails to check the efficacy and impact of the ongoing vaccines. While following the guidelines of wearing masks, social distancing, frequent hand-washing are vital steps to stop the viral transmissions of these variants that can further restrict the chances of virus to mutate.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.