m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1

Available online at www.sciencedirect.com

ScienceDirect

j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / m j a fi

Review Article

SARS-CoV-2: Camazotz's Curse

K.B. Anand a, S. Karade b, S. Sen c,*, R.M. Gupta d

a
Associate Professor, Department of Microbiology, Armed Forces Medical College, Pune 411040, India
b
Assistant Professor, Department of Microbiology, Armed Forces Medical College, Pune 411040, India
c
Professor & Head, Department of Microbiology, Armed Forces Medical College, Pune 411040, India
d
Professor (Microbiology) & Dean and Deputy Commandant, Armed Forces Medical College, Pune 411040, India

article info abstract

Article history: The world is currently face to face with a pandemic which is spreading rapidly across the
Received 17 March 2020 globe caused by SARS-CoV-2, a strain of Coronaviruses (CoVs) belonging to subgenus
Accepted 22 April 2020 Sarbecovirus of genus Betacoronavirus. World Health Organisation (WHO) on 11 Feb 20
Available online 27 April 2020 named this disease caused by SARS-CoV-2 as Covid-19. This pandemic is spreading rapidly
and more than 20,00,000 cases have occurred globally. The human Coronaviruses discov-
Keywords: ered in 1960s were considered potentially harmless endemic viruses with seasonal distri-
Human coronavirus bution before late 2002. The CoVs are found in a large number of domestic and wild
COVID-19 animals and birds. The first pandemic caused by Coronavirus caused by SARS-CoV was
SARS CoV-2 recognized in the late 2002 in Guangdong Province and resulted in widespread morbidity
SARS-CoV and mortality. This was followed by MERS-CoV which began in 2012 in the Arabian
MERS-CoV peninsula with multiple outbreaks related to it in various parts of the globe. Various studies
have suggested how these viruses made their entry from their natural reservoir bats via
intermediate host like civets and camels in case of SARS-CoV and MERS-CoV respectively.
The intermediate host of the SARS-CoV-2 still needs to be established. The SARS-CoV-2 has
96.2% similarity to the bat Severe Acute Respiratory Syndrome related-Coronavirus (SARSr-
CoV RaTG13). SARS-CoV-2 has been found to be more distant in relation to SARS-CoV (79%)
and MERS-CoV (50%). At the whole genome sequence level pangolin CoV and SARSr-CoV
RaTG13 show 91.02% and 96.2% similarity with SARS-CoV-2 but the S1 subunit of spike
protein of pangolin CoV is more closely related to SARS-CoV-2 than SARSr-CoV RaTG13.
The genetic analysis of the currently circulating strains of the pandemic have shown 99.98-
100% similarity in their genomes implying a recent shift to humans. The animal source of
SARS-CoV-2 needs to be identified to implement control measures in the present
pandemic. Also, how the virus moves interspecies will help predict and prevent future
pandemics.
© 2020 Director General, Armed Forces Medical Services. Published by Elsevier, a division of
RELX India Pvt. Ltd. All rights reserved.

* Corresponding author.
E-mail address: [email protected] (S. Sen).
https://doi.org/10.1016/j.mjafi.2020.04.008
0377-1237/© 2020 Director General, Armed Forces Medical Services. Published by Elsevier, a division of RELX India Pvt. Ltd. All rights
reserved.
 m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1 137

CoV and MERS-CoV are the two major causes of severe

Coronaviruses: origin and evolution pneumonia in humans.5,9e11 SARS-CoV 2 is the seventh CoV
known to infect humans.
Introduction
SARS-CoV and MERS-CoV
A pandemic of acute respiratory illness has shocked the
world. Initially reported from Wuhan province, China in Dec SARS was the first known pandemic caused by a CoV. The
2019, currently, the viral illness is rapidly spreading across the disease got recognized in the late 2002 with the outbreak of
globe. The virus spreads by droplet transmission, contact with acute atypical community acquired pneumonia noticed first
infected case or contact with contaminated fomites. The dis- at Guangdong Province and 29 countries got affected by the
ease was first recognized after a cluster of pneumonia spread.5,9,10
outbreak was reported in late December 2019 from Wuhan, The 2003 SARS-CoV pandemic resulted in widespread
China. A new, human coronavirus (HCoV) was isolated from morbidity and mortality and the same ended in Jun 2003.9,10
these cases and identified as a betacoronavirus and provi- This was followed by a novel CoV, which was isolated from
sionally named 2019 novel corona virus (2019-nCoV) using a Saudi Arabian patient with severe acute respiratory syn-
next-generation sequencing technology.1,2 On 11 Feb 2020, the drome in June 2012 and the virus was later named Middle East
International Committee on taxonomy of Viruses named the Respiratory Syndrome - Coronavirus (MERS-CoV). Since then,
virus as “severe acute respiratory syndrome coronavirus 2” multiple outbreaks have been reported in or been epidemio-
(SARS-CoV2) and World Health Organisation announced logically linked to the Arabian Peninsula.5,11
COVID-19, the name of the new disease caused by it. Many Besides SARS-CoV and MERS-CoV, the other human coro-
authors have studied the genome sequences of the circulating naviruses are global in their distribution in a seasonal
virus to understand the viral dynamics and the way this new endemic way and are responsible for less than 0.6e2.5% of
strain has made its way into the human population and lead adult community acquired pneumonia patients.12
to the current pandemic. Speculations considering it a labo-
ratory constructed or bioengineered virus have also emerged. Origin
Studies to suggest that it naturally evolved from its existing
ancestors in zoonotic reservoirs have also been published.3 CoVs have been found in large number of domestic and wild
However the research to establish the real origin of SARS- mammals and birds. There are studies to suggest that birds
CoV2, is still underway. and bats are the natural reservoirs of the virus.2,13,14 Corona-
viruses also have a potential for interspecies transmission
Discovery which can also cause zoonotic outbreaks.11 Studies have
suggested a bat origin of the HCoV-229E and HCoV-NL63.
Coronavirus (CoV) was first isolated in 1965 by Tyrrell et al. HCoV-229E have originated from bats with camels acting as
from the nasal washings of a male child.4 Since their discovery intermediate hosts.13e16 Molecular evolutionary analysis of
in 1965, number of circulating strains of coronaviruses were HCoV-OC43 isolates suggests Bovine CoV (BCoV) is their
identified, which were considered harmless pathogens, genetically closest counterpart compared with other CoV
causing common cold and mild upper respiratory illness.5 species17,18 A high similarity was observed between BCoV,
canine respiratory coronavirus (CRCoV) and human corona-
Structure virus OC43 (HCoV-OC43).12 The evolution of HCoV-OC43 has
been shown to be by recombinant events.12 However the
Coronaviruses (CoVs) have large linear positive stranded RNA origin of CoV HKU1 is currently unknown.18 Phylogenetic
genomes approximately 30 kb in size (26e32 kb) and they are analysis has revealed that 2019-nCoV fell within the subgenus
about 125 nm in diameter,6,7 and comprise four genera (alpha-, Sarbecovirus of the genus Betacoronavirus. The homology
beta-, gamma-, and delta-coronavirus).6e8 The spherical or modelling by the authors derived that 2019-nCoV had a
pleomorphic virions are enveloped and contain a helical similar receptor-binding domain (RBD) structure to that of
nucleocapsid of nucleoproteins (N) associated with the RNA SARS-CoV, despite amino acid variation at some key residues
genome. Embedded in the envelope are 20 nm trimer of spike and the ability of the virus to bind to the Angiotensin con-
glycoprotein (S), also called peplomers which have a club verting enzyme 2 (ACE2) in humans.19
shaped morphology and facilitate attachment to cells. Enve- The genomic characterization of the novel Coronavirus
lope also contains integral membrane (M) and envelope (E) from Wuhan cluster by various study groups was based on
proteins. CoVs belonging to the Beta coronavirus lineage have next generation sequencing of samples from bronchoalveolar
5e7 nm spikes of an additional membrane glycoprotein hem- lavage fluid and cultured isolates. Lu et al. studied the CoV
agglutinin esterase (Fig. 1).7 isolated from nine inpatients, eight of whom had visited the
Huanan seafood market in Wuhan. Their study showed that
Human coronaviruses 2019-nCoV was related (with 88% identity) to two bat-derived
severe acute respiratory syndrome (SARS)-like coronaviruses,
Till now Six human CoVs (HCoVs) have been confirmed:
bat-SL-CoVZC45 (GenBank accession number MG 772933) and
HCoV-NL63 and HCoV-229E, which belong to the alpha-
bat-SL-CoVZXC21 (MG772934) and more distant from SARS-
coronavirus genus; and HCoV-OC43, HCoV-HKU1, SARS-CoV,
CoV (about 79%) and MERS-CoV (about 50%).20 However their
and MERS-CoV, belong to the beta-coronavirus genus. SARS-
study also revealed that S gene of 2019-nCoV had the lowest
138 m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1

Fig. 1 e Structure of SARS-CoV2 virus.

sequence identity with bat-SL-CoVZC45 and bat-SL- (97.5% nucleotide identity), Open Reading Frame 7a (ORF7a)
CoVZXC21, at only around 75%.20 Zhou et al. demonstrated and Open Reading Frame10 (ORF10).22 The S1 protein which
that the novel virus has 96.2% similarity to a bat SARS-related contains the RBD, is phylogenetically closer to pangolin-CoV
Coronavirus (SARSr-CoV; RaTG13 (MN996532.1)).21 Zhang et al. than RaTG13 and this RBD region within the S1 was found to
showed that some pangolin CoV genes show higher amino be conserved between Pangolin CoV and SARS-CoV2. The CoV
acid sequence identity to SARS-CoV-2 than to RaTG13 genes spike (S) protein consisting of 2 subunits (S1 and S2), mediates
which included Open Reading Frame 1b (ORF1b), spike protein infection of receptor-expressing host cells and the similarity

LC528233.1 SARS-CoV-2/Hu/DP/Kng/19-027
MT159710.1 2019-nCoV/USA-CruiseA-9/2020
LC528232.1 SARS-CoV-2/Hu/DP/Kng/19-020
MN908947.3 SARS-CoV-2/Wuhan-Hu-1
NC 045512.2 SARS-CoV 2/Wuhan-Hu-1
100 MT066156.1 SARS-CoV-2/INMI1/human/2020/ITA
SARS-CoV2 group
MT072688.1 SARS0CoV-2/61-TW/human/2020/ NPL
MT012098.1 SARS-CoV-2/29/human/2020/IND
MN975262.1 2019-nCoV HKU-SZ-005b 2020
100
MN938384.1 2019-nCoV HKU-SZ-002a 2020
MT050493.1 SARS-CoV-2/166/human/2020/IND
100
MT106052.1 2019-nCoV/USA-CA7/2020 Beta-CoV

MN996532.1 Bat CoV RaTG13

100
MT084071.1 Pangolin CoV
MG772933.1 SARSr- CoV ZC45
96
100 MG772934.1 Bat SARSr-CoVZXC21
KF294457.1 Bat SARSr CoV Longquan-140
100
100 FJ588686.1 SARS CoV Rs672/2006
100 100 NC 004718.3 SARS CoV
NC 014470.1 Bat CoV BM48-31/BGR/2008
NC 025217.1 Bat Hp-beta Cov/Zhejiang2013
MH734115.1 MERS-CoV camel/Kenya/C1272/2018
100 NC 019843.3 MERS-CoV

100 NC 006213.1 Human CoV OC43

NC 006577.2 Human CoV HKU1
100 NC 005831.2 Human CoV NL63
Alpha-CoV
100 NC 002645.1 Human CoV 229E

0.10

Fig. 2 e Phylogenentic Relationship of CoVs Based on the Whole Genome. Maroon text denotes SARS-CoV2. Pink text
denotes Pangolin CoV. Green text denotes RaTG13. Blue text denotes Bat SARSr- CoV ZC45 and Bat SARSr-CoVZXC21. Light
blue denotes SARS CoV. Purple denotes MERS-CoV.
 m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1 139

Fig. 3 e Multiple sequence alignment of protein sequences of 26 circulating strains ORF 8 showing the leucine at position 84
in 15 out of 26 strains.

between S1 protein of pangolin CoV to SARS-CoV2 points po- the nucleotide sequences establishing a relationship between
tential similarity in their pathogenic properties.22 Though the the currently circulating viruses and implying a recent shift to
origin of the SARS-CoV 2 is still a debatable topic but the humans (Fig. 2).
recognition of the intermediate animal host is the crucial step
in preventing further dissemination, future outbreaks and The L & S type of the circulating SARS-CoV2
blocking the interspecies transmission.
Tang et al. have carried out extensive study on the circulating
A naturally originated virus SARS-CoV2 strains and divided the virus into two major types
of SARS-CoV2 define by two single nucleotide poly morphisms
Outbreak of fatal respiratory illnesses in Wuhan, China lead to (SNPs) that show complete linkage.25 The author analysed 103
speculations that SARS-CoV2 could be laboratory manipu- SARS-CoV-2 virus strains and found that 101 showed com-
lated virus, however a study published in Nature Medicine on plete linkage between the two SNPs: 72 strains exhibited a
17 Mar 20 by Andersen et al. concluded that the SARS-CoV2 is “CT” haplotype (defined as “L” type because T28,144 is in the
not a laboratory constructed or manipulated virus based on codon of Leucine) and 29 strains exhibited a “TC” haplotype
the RBD on the SARS-CoV2.3 SARS-CoV-2 has RBD that has (defined as “S” type because C28,144 is in the codon of Serine)
high affinity to ACE2 from humans, ferrets, cats and other at these two sites. Thus the author proceeded to characterise
species with high receptor homology.21 The receptor binding and categorize the SARS-CoV-2 viruses into two major types,
domain in SARS-CoV2 is different from that of SARS-CoV and with L being the major type (~70%) and S being the minor type
the binding of SARS-CoV2 is not optimal based on computa- (~30%). Multiple sequence alignment in MEGA 7 of amino acid
tional analysis leading to the understanding that there is sequences of ORF 8 protein gene of 26 circulating strains from
another mechanism of binding which has arisen out of nat- various locations around the globe to look for the frequency of
ural selection of the virus in the human or human like ACE2.3 the CT haplotype against the TC haplotype at position 84 was
The other salient finding noted of that study is the presence of carried out. Our findings show that 11 out of 26 randomly
a polybasic cleavage site at the junction of S1 and S2, though selected strains showed Serine and 15 showed Leucine at
the role of this is not well established, may allow better cell to position 84 of ORF 8 (57.69%) (Fig. 3).
cell fusion without affecting viral entry.3,23,24
Why SARS CoV2 pandemic is different from previous CoV
Genetic analysis of circulating strains outbreak

We performed phylogenetic analysis of circulating coronavi- Although both SARS CoV2 and prior SARS-CoV utilize ACE2
ruses strains using maximum likelihood method in MEGA receptors to invade respiratory epithelium, the magnitude of
software. The near full length sequences of currently circu- infections caused by SARS-CoV2 is enormous. We are still
lating strains were randomly selected and downloaded from unable to pinpoint the original reservoir for SARS CoV2. SARS-
GeneBank. Recently isolated SARS-CoV2 from India were also CoV genome was found to be 99.8% similar to that from civet
included in the study (Accession no. MT050493.1 and cats. The similarity of whole genome sequence of SARS CoV2
MT012098.1). All sequences showed ~99.98e100% similarity in to pangolins is only 92% and 96.2% to a bat SARS related
140 m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1

coronavirus (SARSr-CoV; RaTG13) which is insufficient to 3. Andersen KG, Rambaut A, Lipkin WL, Holmes EC, Garry RF.
prove beyond doubt that these are the sources of the virus. The Proximal Origin of SARS-CoV-2. ARTIC Network; 17 February
SARS was a relatively rare disease and at the end of the 2020.
4. Tyrrell DAJ, Bynoe ML. Cultivation of a novel type of common
epidemic more than 8000 cases had occurred from 01Nov 2002
cold virus in organ cultures. Brit. med. J. 1965;1:1467e1470.
to 31 Jul 2003 whereas the ongoing Covid 19 caused by SARS- 5. Yin Y, Wunderink RG. MERS, SARS and other coronaviruses as
CoV2 has already caused more than 20, 00, 000 cases across causes of pneumonia. Respirology. 2018;23:130e137.
the globe in a span of approximately five months.26,27 Though 6. Fehr AR, Perlman S. Coronaviruses: an overview of their
the mode of spread of both the viruses is almost same, SARS replication and pathogenesis. Methods Mol Biol.
COV2 is far more infectious than SARS-CoV and the reason for 2015;1282:1e23.
the same is yet to be established. 7. Gadsby NJ, Templeton KE. coronaviruses. In: Caroll Karen C,
Pfaller MA, landry ML, McAdam AJ, Patel R, et al., eds. Manual
of Clinical Microbiology. 12th ed. vol. 2. Washington DC: ASM
Press; 2019:1606e1624 [Chapter 92].
Conclusion 8. de Groot RJ, Cowley JA, Enjuanes L, et al. Order nidovirales. In:
King AMQ, Adams MJ, Carstens EB, Lefkowitz EJ, eds. Virus
The world today is facing a crisis due to the pandemic caused Taxonomy. Ninth Report of the International Committee on
by SAR-CoV2. Based on the available literature and ongoing Taxonomy of Viruses. Amsterdam: Elsevier Academic Press;
2012:785e795.
research, it is interesting to see that the more virulent forms
9. Drosten C, Gunther S, Preiser W, et al. Identification of a novel
(SARS-CoV,MERS-CoV and SARS-CoV2) get adapted to
coronavirus in patients with severe acute respiratory
humans at some point in time of their evolution and this syndrome. N Engl J Med. 2003;348:1967e1976.
moment is crucial to the transmission of these viruses to 10. Ksiazek TG, Erdman D, Goldsmith CS, et al. A novel
humans. The SARS-CoV 2 appears to have originated in bats coronavirus associated with severe acute respiratory
and the intermediate host could be pangolins, however it is syndrome. N Engl J Med. 2003;348:1953e1966.
difficult at this point in time to confidently determine the 11. Wang Y, Sun J, Zhu A, Zhao J, Zhao J. Current understanding
of middle east respiratory syndrome coronavirus infection in
zoonotic source. Once they enter the human population, it is
human and animal models. J Thorac Dis. 2018;10:S2260eS2271.
only a matter of time after which the morbidity and mortality 12. Oong XY, Ng KT, Takebe Y, et al. Identification and
caused by these viruses reach pandemic levels. The whole evolutionary dynamics of two novel human coronavirus
genome sequence analysis of randomly selected currently OC43 genotypes associated with acute respiratory infections:
circulating human strains of SARS-CoV2 available show phylogenetic, spatiotemporal and transmission network
~99.98e100% similarity suggesting a recent introduction of the analyses. Emerg Microb Infect. 2017 Jan 4;6(1):e3. https://
virus into humans. doi.org/10.1038/emi.2016.132.
13. Li W, Shi Z, Yu M, et al. Bats are natural reservoirs of SARS-
However, within these two types of SARS-CoV-2, L type
like coronaviruses. Science. 2005;310(5748):676e679.
(~70%) and S type (~30%) have been observed by Tang et al. 14. Corman VM, Baldwin HJ, Tateno AF, et al. Evidence for an
The strains in L type is derived from S type. The L type as per ancestral association of human coronavirus 229E with bats. J
the author's conclusion appears to be more virulent and Virol. 2015;89(23):11858e11870.
contagious.25 The human-animal interphase created by either 15. Dijkman R, van der Hoek L. Human coronaviruses 229E and NL63:
encroachment of natural habitat of wild animals, maintaining close yet still so far. J Formos Med Assoc. 2009 Apr;108(4):270e279.
https://doi.org/10.1016/S0929-6646(09)60066-8.
domestic animals which get infected or consumption of ani-
16. Corman VM, Eckerle I, Memish ZA, et al. Link of a ubiquitous
mals which may be harbouring the viruses is the tipping point
human coronavirus to dromedary camels. Proc Natl Acad Sci
as per a large number of studies. It is important to understand Unit States Am. 2016:201604472.
viral dynamics so that future outbreaks can be avoided by 17. Bidokhti MRM, Tråve n M, Krishna NK, et al. Evolutionary
active surveillance for these viruses. Regulations also need to dynamics of bovine coronaviruses: natural selection pattern
be formulated to restrict the domestication as well as con- of the spike gene implies adaptive evolution of the strains. J
sumption of wild animals. Gen Virol. 2013;94:2036e2349.
18. Vijgen L, Keyaerts E, Lemey P, et al. Evolutionary history of
the closely related group 2 coronaviruses: porcine
hemagglutinating encephalomyelitis virus, bovine
Disclosure of competing interest coronavirus, and human coronavirus OC43. J Virol.
2006;80:7270e7274.
The authors have none to declare. 19. Woo PC, Lau SK, Lam CS, et al. Discovery of seven novel
mammalian and avian coronaviruses in the genus
deltacoronavirus supports bat coronaviruses as the gene
references source of alphacoronavirus and betacoronavirus and avian
coronaviruses as the gene source of gammacoronavirus and
deltacoronavirus. J Virol. 2012;86:3995e4008.
20. Lu R, Zhao X, Li J, et al. Genomic characterisation and
1. Tan W, Zhao X, Ma X, et al. A novel coronavirus genome
epidemiology of 2019 novel coronavirus: implications for
identified in a cluster of pneumonia casesdWuhan, China
virus origins and receptor binding. Lancet. 2020. https://
2019e2020. China CDC Weekly. 2020;2:61e62.
doi.org/10.1016/S0140-6736(20)30251-8. Epub 2020/02/03.
2. Zhu N, Zhang D, Wang W, et al. A novel coronavirus from
21. Zhou P, Yang X, Wang X, et al. A pneumonia outbreak
patients with pneumonia in China, 2019. N Engl J Med. 2020
associated with a new coronavirus of probable bat origin.
(published online Jan 24.).
Nature. 2020. https://doi.org/10.1038/s41586-020-2012-7.
 m e d i c a l j o u r n a l a r m e d f o r c e s i n d i a 7 6 ( 2 0 2 0 ) 1 3 6 e1 4 1 141

22. Zhang T, Wu Q, Zhang Z. Probable pangolin origin of SARS- 25. Tang X, Wu C, Li X, et al. On the origin and continuing
CoV-2 associated with the COVID-19 outbreak. Curr Biol. evolution of SARS-CoV-2. Natl Sci Rev. 2020.
March 13 2020. https://doi.org/10.1016/j.cub.2020.03.022. 26. Summary of Probable SARS Cases with Onset of Illness from 1
23. Walls AC, Park YJ, Tortorici MA, Wall A, McGuire AT, November 2002 to 31 July 2003. World Health Organization; 21
Veesler D. Structure, function, and antigenicity of the SARS- April 2004.
CoV-2 spike glycoprotein. Cell. 2020;6(20):30262. S0092e8674. 27. Coronavirus COVID-19 Global Cases by the Center for
24. Nao N, Yamagishi J, Miyamoto H, et al. Genetic predisposition Systems Science and Engineering (CSSE) at Johns Hopkins
to acquire a polybasic cleavage site for highly pathogenic University (JHU). ArcGIS. Johns Hopkins CSSE. Retrieved 15
avian influenza virus hemagglutinin. mBio. 2017;8. April 2020.