Received: 22 June 2022 | Accepted: 8 August 2022 DOI: 10.1002/jmv.28060 RESEARCH ARTICLE Tracing the origin of Severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2): A systematic review and narrative synthesis Nagendra Thakur1 | Sayak Das1 | Swatantra Kumar2 | Vimal K. Maurya2 | Kuldeep Dhama3 | Janusz T. Paweska4 | Ahmed S. Abdel‐Moneim5 | Amita Jain2 | Anil K. Tripathi2 | Bipin Puri2 | Shailendra K. Saxena2 1 Department of Microbiology, School of Life Sciences, Sikkim University, Tadong, Gangtok, India 2 Centre for Advanced Research (CFAR), Faculty of Medicine, King George's Medical University (KGMU), Lucknow, India 3 Division of Pathology, ICAR‐Indian Veterinary Research Institute, Izatnagar, Bareilly, India 4 Centre for Emerging Zoonotic and Parasitic Diseases, National Institute for Communicable Diseases of the National Health Laboratory Service, PB X4, Sandringham‐Johannesburg, South Africa 5 Abstract The aim of the study was to trace and understand the origin of Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) through various available literatures and accessible databases. Although the world enters the third year of the coronavirus disease 2019 pandemic, health and socioeconomic impacts continue to mount, the origin and mechanisms of spill‐over of the SARS‐CoV‐2 into humans remain elusive. Therefore, a systematic review of the literature was performed that showcased the integrated information obtained through manual searches, digital databases (PubMed, CINAHL, and MEDLINE) searches, and searches from legitimate publications (1966–2022), followed by meta‐analysis. Department of Microbiology, College of Medicine, Taif University, Al‐Taif, Saudi Arabia Our systematic analysis data proposed three postulated hypotheses concerning the Correspondence obscure origin (O). Despite the fact that the zoonotic origin for SARS‐CoV‐2 has not Shailendra K. Saxena, Centre for Advanced Research (CFAR), Faculty of Medicine, King George's Medical University (KGMU), Lucknow 226003, India. Email: shailen@kgmcindia.edu origin of the SARS‐CoV‐2, which include zoonotic origin (Z), laboratory origin (L), and been conclusively identified to date, our data suggest a zoonotic origin, in contrast to some alternative concepts, including the probability of a laboratory incident or leak. Our data exhibit that zoonotic origin (Z) has higher evidence‐based support as compared to laboratory origin (L). Importantly, based on all the studies included, we generated the forest plot with 95% confidence intervals (CIs) of the risk ratio estimates. Our meta‐analysis further supports the zoonotic origin of SARS/SARS‐ CoV‐2 in the included studies. KEYWORDS COVID‐19, laboratory incidence, MERS‐CoV, origin, SARS‐CoV, SARS‐CoV‐2, zoonotic 1 | INTRODUCTION human transmission, massive administration of various vaccines has succeeded in decreasing the global death rate. SARS‐CoV‐2 has Severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2) has spread worldwide since it was first discovered in Wuhan, China been responsible for the global coronavirus disease 2019 (COVID‐19) where its source of transmission to humans seems to be traced to a pandemic with at least 426 million cases and 5.89 million deaths seafood wholesale market.2 1 Despite the ongoing emergence of different Previous epidemics caused by other coronaviruses (CoVs), such variants of SARS‐CoV‐2 with increased efficiency for human‐to‐ as the Severe acute respiratory syndrome coronavirus (SARS‐CoV) in reported to date. J Med Virol. 2022;1–14. wileyonlinelibrary.com/journal/jmv © 2022 Wiley Periodicals LLC. | 1 2 | THAKUR 2002 and the Middle East respiratory syndrome coronavirus (MERS‐ 2.3 | ET AL. Forest plot analysis CoV) in 2012, originated from bats and involved intermediate hosts.3 To date, seven human coronaviruses do exist including human A Forest plot was generated between Z and L origin using Cochrane's Coronavirus‐229E (HCoV‐229E), human Coronavirus‐OC43 (HCoV‐ Review Manager (RevMan, version 5.4; Nordic Cochrane Center, OC43), human Coronavirus‐NL63 (HCoV‐NL63), and human Corona- Copenhagen, Denmark). The risk ratio (RR) at 95% confidence virus‐HKU1 (HCoV‐HKU1), SARS‐CoV, MERS‐CoV, and SARS‐CoV‐ interval (CI), was calculated to estimate the ratio of the risk in the 2. The former four coronaviruses are the most predominant types of Z group to the risk in the L group. human coronaviruses that cause the common cold.4 Based on the currently available data, it remains unclear whether the inception of SARS‐CoV‐2 is the result of zoonosis caused by a 3 | RESULTS AND DISCUSS I ON wild viral strain or an accidental escape of experimental strains. It is critical to address this issue to develop preventive and biosafety 3.1 | Historical evidence of coronavirus measures. Indeed, the recent zoonosis can justify the need to obtain samples from natural ecosystems, farms, and breeding facilities to The HCoV‐229E was first discovered in the United Kingdom in 19666 prevent spillover. On the contrary, a laboratory escape would followed by the discovery of HCoV‐OC43 in 1967 from a patient necessitate a thorough re‐evaluation of the risk/benefit balance of with respiratory distress in the United Kingdom.7,8 The HCoV‐NL63 various laboratory methods and the stringent implementation of was isolated during the 2002‐to‐2003 winter season in the biosafety standards. Several theories regarding the origin of SARS‐ Netherlands,9 and HKU1 was first reported in an individual from a CoV‐2 are considered. The critical need to advance biosafety large Chinese metropolis (Shenzhen, Guangdong) who developed standards at all laboratory levels is paramount as experimental pneumonia in the winter of 2004.10,11 The SARS‐CoV was first virology research on dangerous pathogens develops to reduce the detected in November 2002 in Foshan, China.12 It has infected threat of pandemics to the environment and human civilization. several people with 8447 cases and caused 813 deaths (9.6% case Therefore, in the present study, we have performed a systematic fatalities); it was contained in July 2003.13 MERS‐CoV was first review, followed by meta‐analysis to decipher the origin of SARS‐ detected in Saudi Arabia in June 2012; however, neutralizing CoV‐2. antibodies have been detected in archival serum samples from dromedary camels in Somalia and Sudan in 1983.14 MERS‐CoV has been reported in 27 more countries in the Middle East, North Africa, 2 | M E TH O D S Asia, Europe, and the United States15 resulting in more than 2585 cases and 890 deaths (case‐fatality ratio of 34.4%) from the virus. 2.1 | Literature search Saudi Arabia, the United Arab Emirates, and the Republic of Korea were the countries with the most outbreaks.16 In comparison to A systematic review was performed by the sources listed in females, a higher percentage of males (about 63%) were severely Supporting Information: Table S1. The sources used for the analysis affected (approx. 37%). MERS‐CoV cases were recorded from nearly were searches every region of the Middle East countries, while Riyadh (30%) and (2000–2022), and CINAHL searches (2000–2022). The major key- PUBMED searches (1966–2022), MEDLINE Jeddah (29%) alone accounted for nearly two‐thirds of the cases.17 words used for indexing the databases were SARS, SARS‐CoV‐2, Later, in December 2019, SARS‐CoV‐2 has emerged in Wuhan, Hubei COVID‐19, coronaviruses, origin, virus, FCS (furin cleavage sites), province, where cases of severe pneumonia were reported.18,19 On spike proteins, bats, novel, and so forth (Supporting Information: March 11, 2020, SARS‐CoV‐2 was declared as the first ever Table S1). This was followed by elaborative discussions with the coronavirus pandemic. experts. Datasets available from NCBI were used for the authentic HCoV‐229E, HCoV‐NL63, SARS‐CoV, MERS‐CoV, and SARS‐ CoV‐2 originated from ancestral bat CoVs,20‐25 whereas the rodent validation of data. CoVs are the ancestral viruses of both HCoV‐OC43 and HCoV‐ HKU1.22 Camels are the current known intermediate animal host of 2.2 | Clustering and similarity matrix analysis both HCoV‐229E and MERS‐CoV.26,27 Although HCoV‐OC43 showed antigenic similarity to bovine CoV suggests a relatively Year‐wise clustergrams/heatmaps were generated to visualize the recent zoonotic transmission event that dates their most recent origin of SARS‐COV‐2 from different sources (Supporting Informa- common ancestor to around 1890.8 HCoV‐NL63 is assumed to be tion: Table S1) specifically zoonotic origin (Z), laboratory origin (L), evolved by a recombination event of NL63‐like viruses and 229E‐like and obscure origin (O). The rows and columns were hierarchically viruses circulating in bats28 and a spillover from bats to humans is clustered, using a cosine distance and an average linkage method assumed to happen 563 to 822 years ago.23 Meanwhile, both civet 5 where the included studies were clustered in rows. Moreover, we cats and raccoon dogs are possible intermediate hosts to the SARS‐ generated the similarity matrix of these origin sources (Supporting CoV.29,30 Although there is no current confirmed intermediate host Information: Table S1). for the SARS‐CoV‐2, pangolins were considered as the incriminated THAKUR | ET AL. 3 hosts31 while HCoV‐HKU1 has an unknown animal origin.11 SARS‐ formed based on the available evidence and which of the recent CoV‐2 exhibits several hallmarks of previous zoonotic outbreaks. It findings or analysis would provide additional information to trace the bears a striking resemblance to the SARS‐CoV, which infected many origin of SARS‐CoV‐2. The first issue in tracing its origin is individuals in the Foshan (2002) and Guangzhou (2003) regions of identification of primary animal hosts before the virus' transmission China.32‐35 The SARS‐CoV outbreaks in these two regions have to humans. CoVs from chiropterans are often transmitted between resulted in a significant increase in the number of people infected bat species and are occasionally transmitted to other mammals, with the virus. SARS‐CoV‐2 outbreaks have been associated with according to the results of a previous phylogenetic analysis.45 Point exposure to wet animal markets in Wuhan (2019), which may mutations and recombination events, common in coronaviruses, are facilitate the transmission of this virus.32 involved in virus co‐evolution with their hosts and adaptation to new hosts.46 Because mosaicism biases the whole genome‐based phylogenetic inference, the resulting tree would reflect a blend of the 3.2 | Significance of the Spike (S) protein of SARS‐ CoV‐2 diverse developmental pathways pursued by the different open reading frames (ORFs), which poses specific challenges. Hence, it is crucial to recognize the recombinant fragments and make different The S protein is a crucial glycoprotein involved in receptor binding phylogenetic inferences for each of them. SARS‐CoV‐2 is thought to and cell entry. The S protein is cleaved from two locations, S1/S2 result from several recombination events among chiropteran CoVs, and the S2′ site, following receptor engagement to promote virus which are probably the principal reservoir of the virus. Because of its 32 According to preliminary structural studies, critical function in the interaction with the host ACE2 receptor and SARS‐CoV‐2 has a higher affinity for the angiotensin‐converting virus entry, the effect of recombination is very significant for the enzyme‐ 2 (ACE‐2) receptor than the original SARS‐CoV.36‐39 Both adaptability of the S protein.47 Our systematic analysis proposes the COVID‐19 patient sera and monoclonal antibodies (mAbs) against following postulated hypothesis concerning the origin of the SARS‐ the receptor‐binding domain (RBD) had lower results in neutrali- CoV‐2. Bioinformatic studies may further help us to determine the entry into the cell. zation studies, including mutations. 40 These findings indicate the origin of SARS‐CoV‐2. vital function of the furin‐like cleavage site (FCS) in the SARS‐CoV‐ 2 infection, as well as the potential pitfalls of interpreting the results of studies on this virus. The FCS deletion significantly 3.3.1 | Theories of SARS‐CoV‐2 origin affects virus neutralization by the sera collected from COVID‐19 patients by administering specific mAb against the SARS‐CoV‐2 Zoonotic origin (Z) RBD. Although each mAb targets a different location in the RBD, the wild type and mutant type exhibit equal reductions in Bats‐to‐man transmission. Bats were thought to be the original host mAb serum neutralization levels, indicating possible therapeutic when the first genomic material for SARS‐CoV‐2 was available.41 Bat‐ approaches against SARS‐CoV‐2.41 CoV‐RaTG13, a bat coronavirus isolated from Rhinolophus affinis, shares a The S1/S2 furin sensitive proteolytic cleavage site appears to 96% whole‐genome sequence identity with SARS‐CoV‐2. SARS‐CoV‐2 contribute to its infectivity in humans and may be related to its closely related viruses have been found in bats in Southeast Asia, epidemic tendency.42 This insertion is likely new because it is not including China, Thailand, Cambodia, Laos (e.g., BANAL‐52), and found in any viruses related to SARS‐CoV‐2. Like SARS‐CoV‐2, Japan.48,49 However, there is a significant evolutionary gap between HCoVOC43, HCoVHKU1 and MERSCoV possess furin cleavage SARS‐CoV‐2 and the closest related animal viruses. For example, the bat site.20 This finding is significant because this genetic characteristic is virus RaTG13 obtained by the Wuhan Institute of Virology (WIV) has a likely to be involved in bridging the species barrier and increasing the genetic distance of >4% (approximately 1150 mutations) from the SARS‐ efficiency of human‐to‐human transmission, both of which are CoV‐2 Wuhan‐Hu‐1 reference sequence, implying the generations of necessary for an epidemic to occur. Several laboratories are developmental differences31 (Figure 1). Moreover, two studies that conducting and publishing gain‐of‐function (GoF) experiments to analyzed the molecular spectrum of mutations also supported bats‐to‐ explore the association between coronavirus RBD and trans- man direct transmission and disputed the possibility of serial passage in membrane receptors such as ACE2.43 SARS‐CoV‐2 was postulated mouse or human cell lines or chimeric coronaviruses.50,51 Year wise as the outcome of experiments to “humanize” an animal virus of the SARS‐CoV‐2, SARS‐like coronaviruses, and SARS‐CoV‐2 isolates have RaTG13 type,44 but the scientific community has not presented been mentioned in Table 1. persuasive proof to confirm this hypothesis. The widespread genome recombination makes it challenging to determine the viruses that are most similar to SARS‐CoV‐2. Even though the RaTG1318 from the Rhinolophus affinis bat in Yunnan has 3.3 | Origin of SARS‐CoV‐2 the highest average genetic similarity to SARS‐CoV‐2, the historical background of recombination assumes that three other bat viruses, The current understanding of SARS‐CoV‐2 origin is inconclusive. RmYN02, RpYN06, and PrC31, have relatively close viral RNA However, it is useful to consider whether conclusions can already be genome with that of SARS‐CoV‐2 (particularly ORF1ab).52,53 4 | THAKUR ET AL. F I G U R E 1 Phylogenetic of SARS‐like coronaviruses and SARS‐CoV‐2. Phylogenetic relationship showing that the SARS‐CoV‐2 is closely related to the SARS‐like coronaviruses isolated from the bats. However, SARS‐CoV‐2 has been reported in pangolins. Whereas earlier reported SARS‐CoV has been isolated from humans, bats, and civets. SARS‐CoV‐2, Severe acute respiratory syndrome coronavirus 2. Cross‐species transmission is commonly overlooked during its early that may be involved in the potential zoonotic ecological niche phases. It is yet to be identified whether several other human (those circulating in chiropterans and in animals that come into coronaviruses like HCoV‐HKU1 and HCoV‐NL63 have animal origins contact with humans) are being sequenced. Pigs, goats, sheep, cows, or not. Despite the genetic similarity of bat coronaviruses to SARS‐ and cats are examples of mammalian species whose ACE2 receptors CoV is more than 95%, their ability to use hACE‐2 as a receptor might are more similar to the main properties of the human receptor than 54 have taken decades to naturally evolve. those of the chiropterans.56 Construction of pangolin farms and The possibility of direct transmission of bat‐borne coronaviruses intense breeding of minks and raccoon dogs have become to humans seems to be a potential mode of spread. In 2012, many increasingly popular in China, bringing more health concerns in mineworkers were sent to clean bat feces from an abandoned addition to concerns associated with practicality of such domesti- mineshaft in Mojiang. This defunct copper mine in Mojiang, more cation.57 Furthermore, these new alien farms coexist with intense than a thousand miles from Wuhan, is infested with horseshoe bats domestic animal husbandry (such as poultry and pigs), which may (Rhinolophus sinicus), which are the documented hosts of SARS‐like facilitate the development of virus reservoirs (such as influenza) in coronaviruses. Six of these miners contracted a mysterious illness and regions that are densely populated.58 The dependability of the showed symptoms of severe pneumonia and acute respiratory results is determined by the quality of genome sequences, genomic distress syndrome. Three of them died with symptoms suspected restorations, information quality, and integrity of annotations in to be consistent with those of SARS‐like disease. All patients sequence databases.59 exhibited respiratory failure showing interstitial lung disease and Viruses intimately correlated with SARS‐CoV‐2 have been found alveolar lesions. Details about the deaths and symptoms of these in bats and pangolins in Southeast Asia, including China, Thailand, miners were uncovered by a skeptic of the wet‐market hypothesis in Cambodia, and Japan, which have been causing viral infections in the form of a Chinese master's thesis.55 This episode is also referred pangolins for more than 10 years.49 Although viral communication to as the “first episode of the bat coronavirus outbreak” after the was discovered between coronaviruses affecting Malayan pangolins 2002 SARS outbreak. Therefore, it might be postulated that as the (Manis javanica) and those affecting other hosts, it was previously miners were previously working in an environment swarmed with bat thought that pangolin coronavirus had no direct association with feces, and all of the six patients had similar case histories, these SARS‐CoV‐2. Pangolin‐CoV‐2019, a pangolin isolate, only shared a circumstances must have some correlations with the development of 91.02% whole‐genome identity with SARS‐CoV‐2, but higher SARS‐like diseases with pneumonia‐like symptoms or severe sequence homology in the spike glycoprotein (S protein, 97.5%) breathing‐associated symptoms arising from bat feces. A similar coding sequence than Bat‐CoV‐RaTG13.60 As a result, the pangolin is scenario could have happened just before SARS‐CoV‐2. thought to be a possible intermediate host for SARS‐CoV‐2. The RBD Transmission to humans through an intermediate host. The origin of recombination between a virus similar or related to Bat‐CoVRaTG13 SARS‐CoV‐2 was investigated to identify other animal viruses with and a virus similar or related to Pangolin‐CoV‐2019.61 The SARS‐ a high degree of resemblance. As a result, new coronavirus genomes CoV‐2 RBD's binding free energy with human‐ACE2 is significantly of the S protein in SARS‐CoV‐2 is thought to have evolved via the THAKUR | ET AL. TABLE 1 5 Year wise SARS‐CoV‐2, SARS‐like coronaviruses and SARS‐CoV‐2 isolates Virus type Year Sequence ID Accession number Host Country SARS‐CoV 2003 TWS AP006560 Human Taiwan SARS‐CoV 2003 TWH AP006557 Human Taiwan SARS‐CoV 2003 BJ01 AY278488 Human China SARS‐CoV 2003 BJ04 AY279354 Human China SARS‐CoV 2004 Sin846 AY559094 Human Singapore SARS‐CoV 2004 Sin842 AY559081 Human Singapore SARS‐CoV 2005 Sino1_11 AY485277 Human China SARS‐CoV 2005 GZ0401 AY568539 Human China SARS‐CoV 2005 GZ0402 AY613947 Human China SARS‐CoV 2005 Civet020 AY572038 Civet China SARS‐CoV 2005 PC4_227 AY613950 Civet China SARS‐CoV 2005 Civet007 AY572034 Civet China SARS‐CoV 2009 A001 FJ959407 Civet China SARS‐like CoV 2010 HKU3_7 GQ153542 Bat China SARS‐like CoV 2013 RS3367 KC881006 Bat China SARS‐like CoV 2013 WIV1 KF367457 Bat China SARS‐like CoV 2013 RsSHC014 KC881005 Bat China SARS‐like CoV 2013 bat/Yunnan/RaTG13/2013 EPI_ISL_402131 Bat China SARS‐like CoV 2014 LYRa11 KF569996 Bat China SARS‐like CoV 2015 YNLF_34C KP886809 Bat China SARS‐like CoV 2015 bat_SL_CoVZXC21 MG772934 Bat China SARS‐like CoV 2017 RS4231 KY417146 Bat China SARS‐like CoV 2017 RS4084 KY417144 Bat China SARS‐like CoV 2017 Rs9401 KY417152 Bat China SARS‐like CoV 2017 Rs7327 KY417151 Bat China SARS‐like CoV 2017 Rf4092 KY417145 Bat China SARS‐like CoV 2017 Rs4237 KY417147 Bat China SARS‐like CoV 2017 Rs4247 KY417148 Bat China SARS‐like CoV 2017 As6526 KY417142 Bat China SARS‐like CoV 2017 Rs4081 KY417143 Bat China SARS‐like CoV 2017 Rs672 KY417143 Bat China SARS‐like CoV 2017 pangolin/Guangxi/P2V/2017 EPI_ISL_410542 Pangolin China SARS‐like CoV 2017 pangolin/Guangxi/P5E/2017 EPI_ISL_410541 Pangolin China SARS‐like CoV 2017 pangolin/Guangxi/P5L/2017 EPI_ISL_410540 Pangolin China SARS‐like CoV 2017 pangolin/Guangxi/P1E/2017 EPI_ISL_410539 Pangolin China SARS‐like CoV 2017 pangolin/Guangxi/P3B/2017 EPI_ISL_410543 Pangolin China SARS‐like CoV 2017 pangolin/Guangxi/P4L/2017 EPI_ISL_410538 Pangolin China SARS‐like CoV 2017 bat_SL_CoVZC45 MG772933 Bat China SARS‐like CoV 2019 Bat/Yunnan/RmYN01/2019 EPI_ISL_412976 Bat China (Continues) 6 | THAKUR ET AL. (Continued) TABLE 1 Virus type Year Sequence ID Accession number Host Country SARS‐CoV‐2 2019 Wuhan/WIV05/2019 MN996529 Human China SARS‐CoV‐2 2019 Wuhan‐Hu‐1/2020 WH‐Human_1 Human China SARS‐like CoV 2019 bat/Yunnan/RmYN02/2019 EPI_ISL_412977 Bat China SARS‐like CoV 2019 pangolin/Guangdong/P2S/2019 EPI_ISL_410544 Pangolin China SARS‐CoV‐2 2020 Japan/KY‐V‐029/2020 LC522972 Human Japan SARS‐CoV‐2 2020 Sweden/01/2020 MT093571 Human Sweden SARS‐CoV‐2 2020 USA/IL1/2020 MN988713 Human USA SARS‐CoV‐2 2020 Nepal/61/2020 MT072688 Human Nepal SARS‐CoV‐2 2020 USA/CA1/2020 MN994467 Human USA lower than that of SARS, which explains the infectious capacity of 62 These findings are consistent with the emergence of SARS‐CoV‐ Although the potential importance of the RBD 2, which is associated with one or more infected animals, as well as discovered in pangolin CoV‐2 has already been established, the with spillovers from numerous infected or extremely susceptible region of high resemblance between pangolin virus and SARS‐CoV‐2 animals transported into or between Wuhan marketplaces, primarily is short, and the possibility of pangolin‐to‐human transmission could through consensual networks and sold for human consump- be very low. Moreover, even the pangolin viruses most closely tion.18 Similar to SARS‐CoV, which was reported to have high levels SARS‐CoV‐2. related to SARS‐CoV‐2 (such as MP789), including its bat coronavirus of transmission, seroprevalence, and genetic variability in animals in relatives (notably RaTG13 and RmYN02), have a low identity rate with the Dongmen market in Shenzhen and the Xinyuan market in SARS‐CoV‐2, implying that closer relatives and possibly more recent Guangzhou, the virus might have proliferated across several intermediate hosts are still unknown.63 Hence, an in‐depth statistical regions.65 analysis of the genomic recombination across coronaviruses from Chinese authorities have conducted a sero‐prevalence survey of various hosts, particularly between pangolin and bat coronaviruses, SARS‐CoV‐2 among animals during the initial period of the pandemic; should be conducted to trace the origin of SARS‐CoV‐2 and uncover however, they did not find any seropositive animals.52 Apart from evolutionary patterns. these studies, only a few research investigations have been Millions of live wild animals, comprising high‐risk species such as conducted on mammals in the Wuhan or Yunnan region, which civets and raccoon dogs, were sold at Wuhan marketplaces in 2019, suggests the presence of an intermediate host for SARS‐CoV‐2.63 In including the Huanan marketplace. 64 SARS‐CoV‐2 was discovered in the last 2 years after the pandemic began, no intermediate host has samples taken from the Huanan market, primarily in the western been reported or identified. By contrast, the intermediate host of section, which sells wildlife and domestic animal products, as well as SARS and MERS was identified within 6 months. Thus, it remains from the sewage areas.65 Even though animal carcasses tested challenging to confirm the intermediate host 2 years after the negative for SARS‐CoV‐2 retrospectively, they were not the typical outbreak of COVID‐19. Moreover, investigating the marketplace that live animal species usually sold in this type of market and did not is now considered the “first victim of COVID‐19 pandemic” may not include raccoon dogs and other animals that are susceptible to SARS‐ be sufficient to determine the source of the current outbreak. All CoV‐2.64 The earliest split in the SARS‐CoV‐2 phylogeny identified possible traces, such as raw animal products used for trading or two lineages, A and B,45 which apparently spread simultaneously. animal corpses, have been destroyed as preventive measures to Lineage B was observed in individuals exposed to other marketplaces eliminate further spillover chances.66 Thus, in all possibilities, as well as those with later cases in Wuhan and other parts of China, humanity might never know the intermediate host that could whereas lineage A was observed in individuals exposed to other transmit the virus to humans, leading to the outbreak. marketplaces as well as those with later cases in Wuhan and other More recently, SARS‐CoV‐2 B.1.1.52‐infected 19/131 white‐ parts of China.65 The lineage A refers to Wuhan/WH04/2020 tailed deer as evidenced by the presence of neutralizing antibodies (EPI_ISL_406801), sampled on January 5, 2020, that shared two and the presence of viral RNA in one animal. This finding could be nucleotides (positions 8782 in ORF1ab and 28144 in ORF8) with the very helpful in finding potential intermediate hosts. Screening the closest known bat viruses (RaTG13 and RmYN02). Lineage B, referred SARS‐CoV‐2‐specific antibodies to SARS‐CoV‐2 in closely related to those strains that had different nucleotides present at those sites animal species in wet markets in China is highly recommended. Such as observed in Wuhan‐Hu‐1 (GenBank accession no. MN908947) an investigation could help in assessing the possible intermediate sampled on December 26, 2019.45 animal hosts for SARS‐CoV‐2 that might spillback to humans.67 THAKUR | ET AL. 7 applied at the WIV to generate infectious SARS‐CoVs based on Laboratory origin (L) bat sequencing data. Hence, gain‐of‐function studies should Seepage from a laboratory incident. The emergence and human ideally use a known SARS‐CoV genetic backbone or, at the very transmission of SARS‐CoV is an example of a laboratory incident least, a virus that has been identified through sequencing. that resulted in single illnesses and temporary transmission chains. Previous scientific work at the WIV using recombinant corona- Apart from the Marburg virus,68 all pathogens that have escaped the viruses employed a genomic framework (WIV1) unrelated to laboratory setting are easily identifiable viruses capable of human SARS‐CoV‐2, and did not have the genetic markers that would infection and have been linked to long‐term research in slightly be expected from laboratory experiments.79 elevated settings. An example of the globally acknowledged human vi. There is no reasonable rationale for establishing novel genetic epidemic or pandemic resulting from scientific activities is the 1977 engineering approaches using an undocumented virus, consid- A/H1N1 influenza pandemic, which was most likely caused by a ering that there is no evidence or mention of a similar virus‐like large‐scale vaccination challenge trial.69 SARS‐CoV‐2 from WIV or any nearly related candidates other In 2021, all the available literature suggested that the emergence than RaTG13. Hence, it is not reasonable to say that SARS‐CoV‐ of SARS‐CoV‐2 was not due to an accidental escape of a laboratory 2 was present in the laboratory before the pandemic in any strain and most likely had a zoonotic origin.4 The assumptions were laboratory escape scenario; however, there is no factual data to based on the following observations: prove it, and no sequence retrieved that can be referred to as progenitor. i. None of the epidemics were caused by a novel virus escaping vii. One example of a laboratory escape scenario is the accidental from a laboratory; moreover, there is no proof that the WIV infection during the serial passage of SARS‐CoV‐like viruses in conducted any previous research on SARS‐CoV‐2 or that any ordinary laboratory animals such as mice. By contrast, early ancestor virus existed before the COVID‐19 pandemic. Since SARS‐CoV‐2 isolates could not infect wild‐type mice.80 viruses are neutralized during RNA extraction, viral genome Although animal models are useful for studying the course of sequencing performed without cell culture does not pose a risk infection in vivo and testing various vaccines, they typically lead of virus transmission, and this procedure was performed at the to the development of moderate or atypical disease in hACE2 WIV.70 After sequencing the viral samples, no incidences of transgenic mice.81 These findings contradict the fact that a laboratory escape were reported. Reported experimental break- certain virus is chosen for use in animal models due to its outs have been linked to the benchmark cases' job and familial increased pathogenicity and transmissibility to infect susceptible contacts, as well as points of origin.71 rodents’ multiple times. SARS‐CoV‐2 has now been generated82 ii. After a thorough investigation and tracking of early instances of and serially passed into mice,83 although adaptation in mice the COVID‐19 epidemic, none of the episodes have been linked requires specific mutations in the spike protein, such as to the staff working at the WIV laboratory; when tested for N501Y.84 N501Y has appeared convergently in several human 72 SARS‐CoV‐2 in March 2020. Reports of illnesses caused by SARS‐CoV‐2 variants of concern, most likely as a result of the SARS‐CoV‐2 should be validated to confirm if they are caused selection for a higher ACE2‐binding affinity.85 If SARS‐CoV‐2 by the virus during the period of heightened influenza was produced from attempts to adapt a SARS‐CoV to be used in transmission as well as other respiratory virus transmissions.72 animal models, it would have acquired mutations such as N501Y iii. According to the reports of previous studies, the WIV has to allow efficient replication in that model, but there is no successfully isolated three SARS coronaviruses from bats (WIV1, evidence to support that such mutations existed at the WIV16, and Rs4874) and has a vast library of bat‐derived commencement of the outbreak. Given its poor pathogenicity 73,74 Notably, SARS‐CoV is more closely linked to all in commonly employed laboratory animals and the lack of three viruses than SARS‐CoV‐2. However, the RaTG13 virus genomic markers compatible with rodent adaptation, SARS‐ materials. from the WIV has never been isolated or cultivated and only CoV‐2 is unlikely to have been acquired by laboratory employ- exists in the form of a nucleotide sequence derived from short ees during viral pathogenesis or GoF studies. sequencing reads.72 iv. Although no existing evidence shows that the FCS site is Obscure origin (O) artificially inserted in the laboratory, insertion of the FCS and RBD was assumed to be induced by site‐directed mutagenesis.75 Frozen food theory. On February 9, 2020, the World Health However, such speculation was aborted by the fact that a Organization (WHO) and Chinese investigations hypothesized that deletion of FCS did occur by serial passage of SARS‐CoV‐2 SAR‐CoV‐2 might have been transmitted to individuals handling viruses in Vero E6 cells.76–78 As a result, these approaches are frozen foods.86 However, this hypothesis has received several unlikely to produce SARS‐CoV‐2 progenitors with func- criticisms. SARS‐CoV‐2 was initially detected on a cutting board tional FCS. used to handle imported salmon in Beijing's Xinfadi agricultural v. According to undocumented reports, other techniques, such as produce wholesale market on June 12, 2020. Over the next 2 weeks, the discovery of potential reverse genetics systems, were not 256 individuals were infected with SARS‐CoV‐2, of whom 98.8% had 8 | THAKUR ET AL. a history of exposure to the Xinfadi market.65 The genome premise of these studies was to anticipate and prepare for the sequencing of a SARS‐CoV‐2 virus detected in a sample obtained next pandemic (before the COVID‐19 hit). from the Xinfadi market revealed a European coronavirus strain, providing a strong indication that the re‐emergent COVID‐19 cases in Beijing may be due to imported sources rather than a local transmission.65 At least, nine food contamination incidents have been 3.4.2 | Concern about the exact time of viral emergence recorded around the country since the beginning of July 2020, with SARS‐CoV‐2 being found on imported items, predominantly in The wet market cases have been consistently claimed to be the packing materials.87 Nonetheless, none of these have provided for earliest cases of the outbreak, lending credence to the “wet‐market any verifiable facts or presented a focused position on the subject to hypothesis.” Some speculations that hypothesized that SARS‐CoV‐2 date. The WHO, the Centers for Disease Control and Prevention, and could be present before December 2019, which were augmented by the Food and Drug Administration in the United States, as well as post hoc data following analysis, showed that it is likely that the other regional regulatory bodies, have advised that there is no SARS‐CoV‐2 probably introduced before December 2019.43 Re- current evidence showing that the SARS‐CoV‐2 that caused COVID‐ searchers discovered and recovered a deleted set of incomplete 19 can spread through foods and that no specific foods should be SARS‐CoV‐2 sequences from the early Wuhan pandemic. Several withdrawn. inferences can be drawn from the analysis. First, the Huanan Seafood Later, in 2022, Multiple Working Hypothesis (MWH) suggested Market sequences, which were the topic of a joint WHO‐China study, that big natural disasters like earthquakes, hurricanes, typhoons, and may not represent all SARS‐CoV‐2 cases in Wuhan around the initial so forth cause higher deaths in a short period on comparing to deaths phases of the outbreak. According to the lost files and accessible caused by naturally occurring (origin) zoonotic viruses like SARS‐ sequences from Wuhan‐infected patients hospitalized in Guangdong, CoV‐2.88 A natural origin zoonotic virus has a remote possibility (i.e., early Wuhan sequences were more likely to carry the T29095C rare events and low risk) of causing deaths as compared to the origin mutation and were less likely to carry T8782C/C28144T than the of viruses through laboratory has a higher probability of inflicting sequences indicated in the joint WHO‐China report.65 Second, there 88 more deaths. are two credible options for SARS‐CoV‐2 progenitors based on the available evidence. ProCoV‐2 was described,93 while the other was a sequence with three mutations (C8782T, T28144C, and C29095T) 3.4 | Concerns that raised suspicions about the current SARS‐CoV‐2 compared with that of the Wuhan‐Hu‐1 sequence. Importantly, both possible progenitors are three mutations closer to the coronavirus cousins of SARS‐bat CoV‐2 than that of the sequences of viruses The zoonotic jump of coronaviruses to humans occurs frequently isolated from the Huanan Seafood Market. The progenitors of all especially when one encounters a situation against the normal known SARS‐CoV‐2 sequences could still be downstream of the concept of nature. The Asian meat markets are known for their exotic sequence that infected patient zero, based on the transmission trades of poached animals for human consumption. These animals are dynamics of the earliest infections.43 This report was also augmented not normally present in close contact with humans. Accordingly, their by the evidence of circulation of SARS‐CoV‐2 in November 2019 in presence together in close contact with each other and to humans France,94 which confirms that the virus emergence was before constitutes a great potential of virus spill‐over from such animals to November 2019. humans. So, the wet market theory is a logical consequence for the possible emergence of the SARS‐CoV‐2. Meanwhile, many facts raised the suspicion of the world for other scenarios that might be 3.5 | Possibilities of Omicron evolution responsible for the current pandemic. The majority of SARS‐CoV‐2 mutations are repetitive or harmful; however, a handful of them improve viral function. D614G, the first 3.4.1 | Work on chimeric coronaviruses known mutation linked to increased transmissibility, was discovered in early 2020. Since then, the virus has mutated, resulting in new Different chimerics of SARS coronaviruses were created in the mutations and a plethora of varieties. They could modify infectivity, Baric laboratory in the USA as reviewed in, 89 including bat‐SCoV transmissibility, or immune escape depending on the genes impacted 90 BtCoV and the location of the mutations. Because of the protein's function HKU5 with the SARS‐CoV spike (S) glycoprotein,91 and murine in the initial virus–cell contact and because it is the most changeable adapted SARS‐CoV with SHC014 spike bat coronavirus. 92 region in the virus genome, mutations that induce differences in the Efficient replication in both mice and human airway cultures SARS‐CoV‐2 spike protein have been among the most investigated was noted in the latter chimeric virus without the need of any to date. genome with the SARS‐CoV receptor‐binding domain, adaptation. These findings highlight the possible risks from the The severity of the sickness caused by virus variants is construction of chimeric from betacoronavirus. 92 The basic determined by their origin, genetic profile (some common mutations THAKUR ET AL. | 9 in the lineage), and the severity of the disease they cause, which number of mutations. The beginnings of Omicron's proximal origins determines the level of worry.95 New varieties can outcompete have swiftly become a contentious matter of contention in the others in the population if they improve their fitness. The Alpha form scientific and public health realms.97 Many of the mutations found in spread faster than previous generations because it was more Omicron were found in previously sequenced SARS‐CoV‐2 variants transmissible. Beta and Gamma versions have accumulated mutations only infrequently,96,98 leading to three popular interpretations about that allow them to partially evade immune systems and reduce its evolutionary past. The first theory is that Omicron disseminated vaccination effectiveness. Later, the Delta variant, discovered in and circulated in a population with limited viral surveillance and March 2021, proliferated and superseded the other variants, sequencing. Second, Omicron could have evolved in a COVID‐19 becoming the most worrying of all the emerging lineages.96 patient who was chronically infected, such as an immuno- The Omicron type has now spread all over the world and is the compromised person, who provided a good host environment for most common. The SARS‐CoV‐2 Omicron variant was first identified long‐term intra‐host viral adaptation. The third scenario is that in South Africa on November 24, 2021, and was quickly designated Omicron collected mutations in a nonhuman host before transferring as a variant of concern (VOC) by the World Health Organization to humans.99 Omicron could have emerged by virus spillover to an (WHO) due to an increase in cases linked to this variant in South animal host/reservoir such as jumping from humans to mice, gained Africa (i.e., Omicron outbreak). Furthermore, the open reading frame mutations favorable to infect mice, and then reinfection to a human encoding Omicron's spike protein (ORF S) has an unusually high host would have occurred, reflecting an inter‐species evolution F I G U R E 2 Theories of SARS‐CoV‐2 origin. SARS‐CoV‐2 shares sequence similarity with intermediate hosts including Bat‐CoV‐RaTG13, a bat coronavirus isolated from Rhinolophus affinis shares 96% whole‐genome sequence identity with SARS‐CoV‐2. SARS‐CoV‐2 has been shown to originate as a spillover from the infected intermediate hosts. Pangolin‐CoV‐2019, a pangolin isolates shared a higher sequence homology of 97.5% with spike glycoprotein. Similarly, SARS‐CoV‐2 might have spillover from infected live wild/domestic animals, including their products. Due to previous leakages of microorganisms from the laboratory, several theories support and contradict the origin of SARS‐CoV‐2 from laboratory leakage. Recent emergence of newer SARS‐CoV‐2 variants, Omicron is imposing serious concern about its origin which might be the result of inter‐species evolution of SARS‐CoV‐2. 10 | THAKUR ET AL. (human‐mice‐human) as proposed on the basis of the presence of included, we generated the forest plot with 95% CIs of the risk ratio mouse‐adapted mutation sites observed that might have facilitated estimates. Our analysis showed that the black diamond supports adaptation of virus to mouse, 98‐102 Therefore, “One Health” approaches have been suggested to be enhanced under the current scenario of Omicron variant outbreaks103,104 (Figure 2). the zoonotic origin of SARS/SARS‐CoV‐2 in the included studies (1966–2022; Figure 4). However, here in a thorough investigative and systematic approach, we have discussed all the possibilities related to the origin of SARS‐CoV‐2. Debunking misinformation and enhancing awareness 4 | C ONC LUS I ON S AND FU TU RE PERS PE C TI V E S about the necessity of research to determine the origin of pathogens are of utmost importance. The fact that the COVID‐19 pandemic occurred in the same region where the WIV is located, a state‐of‐the‐ Based on our keyword searches in PubMed, CINAHL, and MEDLINE art virology laboratory that performs research on bat coronaviruses, library databases, most of the authors favors the zoonotic spillover fueled speculation that SARS‐CoV‐2 was developed in a laboratory. as the most probable origin of SARS‐CoV‐2 whereas origin based on Notwithstanding the rhetoric, there seems to be no compelling proof laboratory spillover is unlikely as no concrete evidence is being that SARS‐CoV‐2 was ever reported to virologists before it emerged shown to cite (Supporting Information: Table S1). (Supporting in December 2019, and all indicators imply that, like SARS and MERS, Information: Table S1 suggests that zoonotic origin (Z) have higher this virus most likely evolved in a bat host unless an unknown human evidence‐based support as compared to laboratory origin (L). This spillover event occurred. has been represented by the heatmap supporting the zoonotic Nevertheless, this accomplished hardly anything to halt the origin of SARS/SARS‐CoV‐2 (Figure 3A). Moreover, the row proliferation of often paradoxical and, at times, completely absurd similarity matrix analysis further supports the zoonotic origin of conspiracy theories that propagated more rapidly than the disease SARS/SARS‐CoV‐2 (Figure 3B). Importantly, based on all the studies outbreak itself. For example, it has been claimed that SARS‐CoV‐2 F I G U R E 3 Heatmap and similarity matrix of SARS‐CoV‐2 origin. (A) Year‐wise studies (Supporting Information: Table S1) supporting the zoonotic origin (Z) of SARS‐CoV‐2 versus laboratory origin (L) versus obscure origin (O). The rows and columns have been hierarchically clustered using cosine‐distance and average linkage, where studies are clustered in rows. Red/blue cells in the matrix represent positive/ negative values in the matrix. (B) Heatmap is showing the row similarity matrix among Z, L, and O. The cells in the matrix represent the similarity between rows, where red/blue represents a positive/negative similarity (measured as 1 − cosine‐distance). THAKUR | ET AL. 11 F I G U R E 4 Forest plot of theories showing the hypothesis of SARS‐CoV‐2 origin. The horizontal line represents the risk ratio estimates at 95% confidence intervals (95% CI). The black diamond supports the zoonotic origin (Z) of SARS‐CoV‐2 based on the included studies (Supporting Information: Table S1). was either the consequence of a laboratory error or was purposefully manufactured or it was produced for GoF investigations, which were previously undertaken with bat SARS‐like coronaviruses to investigate the cross‐species transmission risk. However, performing such RE F ER EN CES 1. 2. research under global prying eyes seems unlikely. Furthermore, disease emergence due to a natural cause has a long history: most 3. Padhi A, Kumar S, Gupta E, Saxena SK. Laboratory diagnosis of novel coronavirus disease 2019 (COVID‐19) infection. In: Saxena S. K., ed. Coronavirus Disease 2019 (COVID‐19): Epidemiology, Pathogenesis, Diagnosis, and Therapeutics. Springer; 2020:95‐107. doi:10. 1007/978-981-15-4814-7_9 4. Holmes EC, Goldstein SA, Rasmussen AL, et al. The origins of SARS‐CoV‐2: a critical review. Cell. 2021;184(19):4848‐4856. doi:10.1016/j.cell.2021.08.017 5. Fernandez NF, Gundersen GW, Rahman A, et al. Clustergrammer, a web‐based heatmap visualization and analysis tool for high‐ dimensional biological data. Sci Data. 2017;4:170151. doi:10. 1038/sdata.2017.151 6. Hamre D, Procknow JJ. A new virus isolated from the human respiratory tract. Proc Soc Exp Biol Med. 1966;121(1):190‐193. doi:10.3181/00379727-121-30734 7. McIntosh K, Dees JH, Becker WB, Kapikian AZ, Chanock RM. Recovery in tracheal organ cultures of novel viruses from patients with respiratory disease. Proc Natl Acad Sci USA. 1967;57(4): 933‐940. doi:10.1073/pnas.57.4.933 Vijgen L, Keyaerts E, Moës E, et al. Complete genomic sequence of human coronavirus OC43: molecular clock analysis suggests a relatively recent zoonotic coronavirus transmission event. J Virol. 2005;79(3):1595‐1604. doi:10.1128/JVI.79.3.1595-1604.2005 van der Hoek L, Pyrc K, Jebbink MF, et al. Identification of a new human coronavirus. Nat Med. 2004;10(4):368‐373. doi:10.1038/ nm1024 Cherry JD, Krogstad P. SARS: the first pandemic of the 21st century. Pediatr Res. 2004;56(1):1‐5. doi:10.1203/01.PDR. 0000129184.87042.FC Woo PC, Lau SK, Chu CM, et al. Characterization and complete genome sequence of a novel coronavirus, coronavirus HKU1, from patients with pneumonia. J Virol. 2005;79(2):884‐895. doi:10. 1128/JVI.79.2.884-895.2005 Chan JF, Kok KH, Zhu Z, et al. Genomic characterization of the 2019 novel human‐pathogenic coronavirus isolated from a patient with atypical pneumonia after visiting Wuhan [published correction appears in Emerg Microbes Infect. 2020;9(1):540]. Emerg Microbes Infect. 2020;9(1):221‐236. doi:10.1080/22221751.2020.1719902 Cleri DJ, Ricketti AJ, Vernaleo JR. Severe acute respiratory syndrome (SARS). Infect Dis Clin North Am. 2010;24(1):175‐202. doi:10.1016/j.idc.2009.10.005 Müller MA, Corman VM, Jores J, et al. MERS coronavirus neutralizing antibodies in camels, Eastern Africa, 1983‐1997. Emerg Infect Dis. 2014;20(12):2093‐2095. doi:10.3201/eid2012. 141026 new viruses that have caused epidemics or pandemics in humans have originated organically from wildlife reservoirs. As a result, the overwhelming opinion is that this virus entered into a susceptible human host through contact with an infected animal, alternatively through contact with infectious animal tissues. A U T H O R C O N TR I B U T I O N S Shailendra K. Saxena conceived the idea and planned the study. Nagendra Thakur, Sayak Das, Swatantra Kumar, Vimal K Maurya, and Shailendra K. Saxena collected the data, devised the initial draft, reviewed the final draft, and contributed equally to this study as the first author. Shailendra K. Saxena, Nagendra Thakur, Sayak Das, Swatantra Kumar, Vimal K. Maurya, Kuldeep Dhama, Janusz T. Paweska, Ahmed S. Abdel‐Moneim, Amita Jain, Anil K. Tripathi, and Bipin Puri finalized the draft for submission. All authors read and approved the final version of the manuscript. 8. A C KN O W L E D G M E N T S The authors are grateful to the Vice Chancellor, King George's 9. Medical University (KGMU) Lucknow, for the encouragement for this work. Ahmed S. Abdel‐Moneim also acknowledges the support of Taif University Researchers Supporting Project No. TURSP‐2020/11. 10. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or 11. financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. 12. CO NFL I CT OF INTERES T The authors declare no conflict of interest. D A TA A V A I L A B I L I T Y S T A T E M E N T 13. The authors confirm that the data supporting the findings of this study are available within the article. ORCID Shailendra K. Saxena http://orcid.org/0000-0003-2856-4185 World Health Organization (WHO). WHO Coronavirus (COVID‐19) Dashboard. Accessed February 28, 2022. https://covid19.who.int/ Harvey WT, Carabelli AM, Jackson B, et al. SARS‐CoV‐2 variants, spike mutations and immune escape. Nat Rev Microbiol. 2021;19(7): 409‐424. doi:10.1038/s41579-021-00573-0 14. 12 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. | THAKUR World Health Organization. Middle East respiratory syndrome coronavirus (MERS‐CoV). 2019. Accessed February 26, 2022. http://www.who.int/emergencies/mers-cov/en/ Lessler J, Salje H, Van Kerkhove MD, et al. Estimating the severity and subclinical burden of Middle East respiratory syndrome coronavirus infection in the Kingdom of Saudi Arabia. Am J Epidemiol. 2016;183(7):657‐663. doi:10.1093/aje/kwv452 Al‐Raddadi RM, Shabouni OI, Alraddadi ZM, et al. Burden of Middle East respiratory syndrome coronavirus infection in Saudi Arabia. J Infect Public Health. 2020;13(5):692‐696. doi:10.1016/j.jiph. 2019.11.016 Zhou P, Yang XL, Wang XG, et al. A pneumonia outbreak associated with a new coronavirus of probable bat origin. Nature. 2020;579(7798):270‐273. doi:10.1038/s41586-020-2012-7 Zhou H, Chen X, Hu T, et al. A novel bat Coronavirus closely related to SARS‐CoV‐2 contains natural insertions at the S1/S2 cleavage site of the spike protein. Curr Biol. 2020;30(19):3896. doi:10.1016/ j.cub.2020.09.030 Abdel‐Moneim AS, Abdelwhab EM. Evidence for SARS‐CoV‐2 infection of animal hosts. Pathogens. 2020;9(7):529. doi:10.3390/ pathogens9070529 Annan A, Baldwin HJ, Corman VM, et al. Human betacoronavirus 2c EMC/2012‐related viruses in bats, Ghana and Europe. Emerg Infect Dis. 2013;19(3):456‐459. doi:10.3201/eid1903.121503 Cui J, Li F, Shi ZL. Origin and evolution of pathogenic coronaviruses. Nat Rev Microbiol. 2019;17(3):181‐192. doi:10.1038/ s41579-018-0118-9 Huynh J, Li S, Yount B, et al. Evidence supporting a zoonotic origin of human coronavirus strain NL63. J Virol. 2012;86(23): 12816‐12825. doi:10.1128/JVI.00906-12 Lau SKP, Woo PCY, Li KSM, et al. Severe acute respiratory syndrome coronavirus‐like virus in Chinese horseshoe bats. Proc Natl AcadSci USA. 2005;102(39):14040‐14045. doi:10.1073/pnas. 0506735102 Pfefferle S, Oppong S, Drexler JF, et al. Distant relatives of severe acute respiratory syndrome coronavirus and close relatives of human coronavirus 229E in bats, Ghana. Emerg Infect Dis. 2009;15(9):1377‐1384. doi:10.3201/eid1509.090224 Perera RA, Wang P, Gomaa MR, et al. Seroepidemiology for MERS coronavirus using microneutralisation and pseudoparticle virus neutralisation assays reveal a high prevalence of antibody in dromedary camels in Egypt, June 2013. Euro Surveill. doi:10.2807/1560-7917.es2013.18.36. 2013;18(36):pii=20574. 20574 Reusken CB, Haagmans BL, Müller MA, et al. Middle East respiratory syndrome coronavirus neutralising serum antibodies in dromedary camels: a comparative serological study. Lancet Infect Dis. 2013;13(10):859‐866. doi:10.1016/S1473-3099(13)70164-6 Tao Y, Shi M, Chommanard C, et al. Surveillance of bat coronaviruses in Kenya identifies relatives of human coronaviruses NL63 and 229E and their recombination history. J Virol. 2017;91(5):e01953‐16. doi:10.1128/JVI.01953-16 Guan Y, Zheng BJ, He YQ, et al. Isolation and characterization of viruses related to the SARS coronavirus from animals in southern China. Science. 2003;302(5643):276‐278. doi:10.1126/science. 1087139 Wang LF, Eaton BTBats. Civets and the emergence of SARS. Curr Top Microbiol Immunol. 2007;315:325‐344. doi:10.1007/978-3540-70962-6_13 Boni MF, Lemey P, Jiang X, et al. Evolutionary origins of the SARS‐ CoV‐2 sarbecovirus lineage responsible for the COVID‐19 pandemic. Nat Microbiol. 2020;5(11):1408‐1417. doi:10.1038/s41564020-0771-4 Saxena SK, Kumar S, Maurya VK, Sharma R, Dandu HR, Bhatt M. Current insight into the novel coronavirus disease 2019 33. 34. 35. 36. 37. 38. 39. 40. 41. 42. 43. 44. 45. 46. 47. 48. 49. ET AL. (COVID‐19). In: Saxena SK, ed. Coronavirus Disease 2019 (COVID‐ 19): Epidemiology, Pathogenesis, Diagnosis, and Therapeutics. Springer; 2020:1‐8. doi:10.1007/978-981-15-4814-7_1 Acute respiratory syndrome. China, Hong Kong special administrative region of China, and Viet Nam. Wkly Epidemiol Rec. 2003;78(11):73‐74. Muller MP, McGeer A. Severe acute respiratory syndrome (SARS) coronavirus. SeminRespirCrit Care Med. 2007;28(2):201‐212. doi:10.1055/s-2007-976492 Poh Ng LF. The virus that changed my world. PLoS Biol. 2003;1(3): E66. doi:10.1371/journal.pbio.0000066 Kumar S, Nyodu R, Maurya VK, Saxena SK. Morphology, genome organization, replication, and pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2). In: Saxena SK, ed. Coronavirus Disease 2019 (COVID‐19): Epidemiology, Pathogenesis, Diagnosis, and Therapeutics. Springer; 2020:23‐31. doi:10. 1007/978-981-15-4814-7_3 Yadav T, Srivastava N, Mishra G, et al. Recombinant vaccines for COVID‐19. Hum Vaccin Immunother. 2020;16(12):2905‐2912. doi:10.1080/21645515.2020.1820808 Kumar S, Maurya VK, Prasad AK, Bhatt MLB, Saxena SK. Structural, glycosylation and antigenic variation between 2019 novel coronavirus (2019‐nCoV) and SARS coronavirus (SARS‐CoV. Virus Dis. 2020;31(1):13‐21. doi:10.1007/s13337-020-00571-5 Kumar S, Nyodu R, Maurya VK, Saxena SK. Host immune response and immunobiology of human SARS‐CoV‐2 infection. In: Saxena SK, ed. Coronavirus Disease 2019 (COVID‐19). Medical Virology: From Pathogenesis to Disease Control. Springer; 2020. doi:10.1007/978-981-154814-7_5 Gupta A, Pradhan A, Maurya VK, et al. Therapeutic approaches for SARS‐CoV‐2 infection. Methods. 2021;195:29‐43. doi:10.1016/j. ymeth.2021.04.026 Malaiyan J, Arumugam S, Mohan K, GomathiRadhakrishnan G. An update on the origin of SARS‐CoV‐2: despite closest identity, bat (RaTG13) and pangolin derived coronaviruses varied in the critical binding site and O‐linked glycan residues. J Med Virol. 2021;93(1): 499‐505. doi:10.1002/jmv.26261 Johnson BA, Xie X, Kalveram B, et al. Furin cleavage site is key to SARS‐CoV‐2 pathogenesis. Preprint. bioRxiv. 2020;2020.08.26. 268854. doi:10.1101/2020.08.26.268854 Bloom JD. Recovery of deleted deep sequencing data sheds more light on the early Wuhan SARS‐CoV‐2 epidemic. Mol Biol Evol. 2021;38(12):5211‐5224. doi:10.1093/molbev/msab246 Kaina B. On the origin of SARS‐CoV‐2: did cell culture experiments lead to increased virulence of the progenitor virus for humans? In Vivo. 2021;35(3):1313‐1326. doi:10.21873/invivo.12384 Rambaut A, Holmes EC, O'Toole Á, et al. Addendum: A dynamic nomenclature proposal for SARS‐CoV‐2 lineages to assist genomic epidemiology. Nat Microbiol. 2021;6(3):415. doi:10.1038/s41564021-00872-5 Tabibzadeh A, Esghaei M, Soltani S, et al. Evolutionary study of COVID‐ 19, severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) as an emerging coronavirus: phylogenetic analysis and literature review. Vet Med Sci. 2021;7(2):559‐571. doi:10.1002/vms3.394 Ahmadi K, Zahedifard F, Mafakher L, et al. Active site‐based analysis of structural proteins for drug targets in different human Coronaviruses. ChemBiol Drug Des. 2021;99:585‐602. doi:10. 1111/cbdd.14004 Temmam S, Vongphayloth K, Baquero E, et al. Bat coronaviruses related to SARS‐CoV‐2 and infectious for human cells. Nature. 2022;604:330‐336. doi:10.1038/s41586-022-04532-4 Wacharapluesadee S, Tan CW, Maneeorn P, et al. Evidence for SARS‐CoV‐2 related coronaviruses circulating in bats and pangolins in Southeast Asia 2021;12(1):1430]. Nat Commun. 2021;12(1): 972. Published 2021 Feb 9. doi:10.1038/s41467-021-21240-1 THAKUR 50. 51. 52. 53. 54. 55. 56. 57. 58. 59. 60. 61. 62. 63. 64. 65. 66. 67. | ET AL. Deng S, Xing K, He X. Mutation signatures inform the natural host of SARS‐CoV‐2. Natl Sci Rev. 2021;9(2):nwab220. doi:10.1093/ nsr/nwab220 Shan KJ, Wei C, Wang Y, Huan Q, Qian W. Host‐specific asymmetric accumulation of mutation types reveals that the origin of SARS‐CoV‐2 is consistent with a natural process. Innovation. 2021;2(4):100159. doi:10.1016/j.xinn.2021.100159 Li Z, Guan X, Mao N, et al. Antibody seroprevalence in the epicenter Wuhan, Hubei, and six selected provinces after containment of the first epidemic wave of COVID‐19 in China. Lancet Reg Health West Pac. 2021;8:100094. doi:10.1016/j.lanwpc.2021. 100094 Li L, Wang J, Ma X, et al. A novel SARS‐CoV‐2 related coronavirus with complex recombination isolated from bats in Yunnan province, China. Emerg Microbes Infect. 2021;10(1):1683‐1690. doi:10.1080/ 22221751.2021.1964925 Hu B, Zeng LP, Yang XL, et al. Discovery of a rich gene pool of bat SARS‐related coronaviruses provides new insights into the origin of SARS coronavirus. PLoS Pathog. 2017;13(11):e1006698. doi:10. 1371/journal.ppat.1006698 Xu L. The Analysis of Six Patients with Severe Pneumonia Caused by Unknown Viruses. Master's Thesis, School of Clinical Medicine, Kun Ming Medical University; 2013. Zhai X, Sun J, Yan Z, et al. Comparison of severe acute respiratory syndrome coronavirus 2 spike protein binding to ACE2 receptors from human, pets, farm animals, and putative intermediate hosts. J Virol. 2020;94(15):e00831‐20. doi:10.1128/JVI.00831-20 Hua L, Gong S, Wang F, et al. Captive breeding of pangolins: current status, problems and future prospects. Zookeys. 2015;507(507):99‐114. doi:10.3897/zookeys.507.6970 Gibbs AJ, Armstrong JS, Downie JC. From where did the 2009 ‘swine‐origin’ influenza A virus (H1N1) emerge? Virol J. 2009;6:207. doi:10.1186/1743-422X-6-207 Sallard E, Halloy J, Casane D, Decroly E, van Helden J. Tracing the origins of SARS‐COV‐2 in coronavirus phylogenies: a review. Environ Chem Lett. 2021;19:1‐17. doi:10.1007/s10311-02001151-1 Lopes LR, de MattosCardillo G, Paiva PB. Molecular evolution and phylogenetic analysis of SARS‐CoV‐2 and hosts ACE2 protein suggest Malayan pangolin as intermediary host. Braz J Microbiol. 2020;51(4):1593‐1599. doi:10.1007/s42770-020-00321-1 Xiao K, Zhai J, Feng Y, et al. Isolation of SARS‐CoV‐2‐related coronavirus from Malayan pangolins. Nature. 2020;583(7815): 286‐289. doi:10.1038/s41586-020-2313-x Zhang T, Wu Q, Zhang Z. Probable Pangolin origin of SARS‐CoV‐2 associated with the COVID‐19 outbreak. Curr Biol. 2020;30(7): 1346‐1351.e2. doi:10.1016/j.cub.2020.03.022 Liu P, Jiang JZ, Wan XF, et al. Are pangolins the intermediate host of the 2019 novel coronavirus (SARS‐CoV‐2). PLoSPathog. 2020;16(5):e1008421. doi:10.1371/journal.ppat.1008421. Xiao X, Newman C, Buesching CD, Macdonald DW, Zhou ZM Animal sales from Wuhan wet markets immediately prior to the COVID‐19 pandemic. Sci Rep. 2021;11(1):11898. doi:10.1038/ s41598-021-91470-2 World Health Organization (WHO)WHO‐convened global study of origins of SARS‐CoV‐2: China Part. 2021. Accessed February 26, https://www.who.int/publications/i/item/whoconvened2022. global-study-of-origins-of-sars-cov-2-china-part Plowright RK, Eby P, Hudson PJ, et al. Ecological dynamics of emerging bat virus spillover. Proc Biol Sci. 2015;282(1798): 20142124. doi:10.1098/rspb.2014.2124 Vandegrift KJ, Yon M, Surendran‐Nair M, et alDetection of SARS‐ CoV‐2 Omicron variant (B.1.1.529) infection of white‐tailed deer. Preprint. bioRxiv. 2022;2022.02.04.479189. doi:10.1101/2022.02. 04.479189 68. 69. 70. 71. 72. 73. 74. 75. 76. 77. 78. 79. 80. 81. 82. 83. 84. 85. 86. 13 Ristanović ES, Kokoškov NS, Crozier I, Kuhn JH, Gligić AS. A forgotten episode of marburg virus disease: Belgrade, Yugoslavia, 1967. Microbiol Mol Biol Rev. 2020;84(2):e00095‐19. doi:10.1128/ MMBR.00095-19 Rozo M, Gronvall GK. The Reemergent 1977 H1N1 Strain and the Gain‐of‐Function Debate. mBio. 2015;6(4):e01013‐15. doi:10. 1128/mBio.01013-15 Blow JA, Dohm DJ, Negley DL, Mores CN. Virus inactivation by nucleic acid extraction reagents. J Virol Methods. 2004;119(2): 195‐198. doi:10.1016/j.jviromet.2004.03.015 Lim PL, Kurup A, Gopalakrishna G, et al. Laboratory‐acquired severe acute respiratory syndrome. N Engl J Med. 2004;350(17): 1740‐1745. doi:10.1056/NEJMoa032565 Cohen J. Wuhan coronavirus hunter Shi Zhengli speaks out. Science. 2020;369(6503):487‐488. doi:10.1126/science.369.6503.487 Liu M, Deng L, Wang D, Jiang T. Influenza activity during the outbreak of coronavirus disease 2019 in Chinese mainland. Biosaf Health. 2020;2(4):206‐209. doi:10.1016/j.bsheal.2020.08.005 Latinne A, Hu B, Olival KJ, et al. Origin and cross‐species transmission of bat coronaviruses in China. Preprint. bioRxiv. 2020. doi:10.1101/2020.05.31.116061 Segreto R, Deigin Y. The genetic structure of SARS‐CoV‐2 does not rule out a laboratory origin: SARS‐COV‐2 chimeric structure and furin cleavage site might be the result of genetic manipulation. BioEssays. 2021;43(3):e2000240. doi:10.1002/bies.202000240. Davidson AD, Williamson MK, Lewis S, et al. Characterisation of the transcriptome and proteome of SARS‐CoV‐2 reveals a cell passage induced in‐frame deletion of the furin‐like cleavage site from the spike glycoprotein. Genome Med. 2020;12(1):68. doi:10. 1186/s13073-020-00763-0 Kumar S, Saxena SK. Structural and molecular perspectives of SARS‐CoV‐2. Methods. 2021;195:23‐28. doi:10.1016/j.ymeth. 2021.03.007 Ogando NS, Dalebout TJ, Zevenhoven‐Dobbe JC, et al. SARS‐ coronavirus‐2 replication in vero E6 cells: replication kinetics, rapid adaptation and cytopathology. J Gen Virol. 2020;101(9):925‐940. doi:10.1099/jgv.0.001453 Andersen KG, Rambaut A, Lipkin WI, Holmes EC, Garry RF. The proximal origin of SARS‐CoV‐2. Nat Med. 2020;26(4):450‐452. doi:10.1038/s41591-020-0820-9 Wan Y, Shang J, Graham R, Baric RS, Li F. Receptor recognition by the novel coronavirus from wuhan: an analysis based on decade‐ long structural studies of SARS coronavirus. J Virol. 2020;94(7): e00127‐20. doi:10.1128/JVI.00127-20 Rathnasinghe R, Strohmeier S, Amanat F, et al. Comparison of transgenic and adenovirus hACE2 mouse models for SARS‐CoV‐2 infection. Emerg Microbes Infect. 2020;9(1):2433‐2445. doi:10. 1080/22221751.2020.1838955 Dinnon KH, 3rd, Leist SR, Schäfer A, et al. A mouse‐adapted model of SARS‐CoV‐2 to test COVID‐19 countermeasures [published correction appears in Nature. 2021;590(7844):E22]. Nature. 2020;586(7830):560‐566. doi:10.1038/s41586-020-2708-8 Leist SR, Dinnon KH, 3rd, Schäfer A, et al. A mouse‐adapted SARS‐ CoV‐2 induces acute lung injury and mortality in standard laboratory mice. Cell. 2020;183(4):1070‐1085.e12. doi:10.1016/j. cell.2020.09.050 Gu H, Chen Q, Yang G, et al. Adaptation of SARS‐CoV‐2 in BALB/c mice for testing vaccine efficacy. Science. 2020;369(6511): 1603‐1607. doi:10.1126/science.abc4730 Khan A, Zia T, Suleman M, et al. Higher infectivity of the SARS‐ CoV‐2 new variants is associated with K417N/T, E484K, and N501Y mutants: an insight from structural data. J Cell Physiol. 2021;236(10):7045‐7057. doi:10.1002/jcp.30367 Liu P, Yang M, Zhao X, et al. Cold‐chain transportation in the frozen food industry may have caused a recurrence of COVID‐19 cases in 14 87. 88. 89. 90. 91. 92. 93. 94. 95. 96. 97. | THAKUR destination: successful isolation of SARS‐CoV‐2 virus from the imported frozen cod package surface. BiosafHealth. 2020;2(4): 199‐201. doi:10.1016/j.bsheal.2020.11.003 Chi Y, Wang Q, Chen G, Zheng S. The long‐term presence of SARS‐ CoV‐2 on cold‐chain food packaging surfaces indicates a new COVID‐19 winter outbreak: a mini review. Front Public Health. 2021;9:650493. doi:10.3389/fpubh.2021.650493 Coccia M. Meta‐analysis to explain unknown cases of the origins of SARS‐COV‐2. Environ Res. 2022;211:113062. doi:10.1016/j. envres.2022.113062 Abdel‐Moneim AS, Abdelwhab EM, Memish ZA. Insights into SARS‐CoV‐2 evolution, potential antivirals, and vaccines. Virology.2021. 2021;558:1‐12. doi:10.1016/j.virol.2021.02.007 Becker MM, Graham RL, Donaldson EF, et al. Synthetic recombinant bat SARS‐like coronavirus is infectious in cultured cells and in mice. Proc Natl Acad Sci USA. 2008;105(50):19944‐19949. doi:10. 1073/pnas.0808116105 Agnihothram S, Yount BL Jr., Donaldson EF, et al. A mouse model for Betacoronavirus subgroup 2c using a bat coronavirus strain HKU5 variant. mBio. 2014;5(2):e00047‐e14. doi:10.1128/mBio. 00047-14 Menachery VD, Yount BL Jr, Debbink K, et al. Author Correction: a SARS‐like cluster of circulating bat coronaviruses shows potential for human emergence. Nat Med. 2020;26(7):1146. doi:10.1038/ s41591-020-0924-2 Kumar S, Tao Q, Weaver S, et al. An evolutionary portrait of the progenitor SARS‐CoV‐2 and its dominant offshoots in COVID‐19 pandemic. Mol Biol Evol. 2021;38(8):3046‐3059. doi:10.1093/ molbev/msab118 Carrat F, Figoni J, Henny J, et al. Evidence of early circulation of SARS‐CoV‐2 in France: findings from the population‐based “CONSTANCES” cohort. Eur J Epidemiol. 2021;36(2):219‐222. doi:10.1007/s10654-020-00716-2 Perez‐Gomez R. The development of SARS‐CoV‐2 variants: the gene makes the disease. J Dev Biol. 2021;9(4):58. doi:10.3390/ jdb9040058 Mistry P, Barmania F, Mellet J, et al. SARS‐CoV‐2 variants, vaccines, and host immunity. Front Immunol. 2022;12:809244. doi:10.3389/fimmu.2021.809244 Sun Y, Lin W, Dong W, Xu J. Origin and evolutionary analysis of the SARS‐CoV‐2 Omicron variant. J Biosaf Biosecur. 2022;4(1):33‐37. doi:10.1016/j.jobb.2021.12.001 98. 99. 100. 101. 102. 103. 104. ET AL. Ma W, Yang J, Fu H, et al. Genomic perspectives on the emerging SARS‐CoV‐2 omicron variant. Genomics Proteomics Bioinformatics. 2022:S1672‐0229. doi:10.1016/j.gpb.2022.01.001 Wei C, Shan KJ, Wang W, Zhang S, Huan Q, Qian W. Evidence for a mouse origin of the SARS‐CoV‐2 omicron variant. J Genet Genomics. 2021;48(12):1111‐1121. doi:10.1016/j.jgg.2021.12.003 Du P, Gao GF, Wang Q. The mysterious origins of the Omicron variant of SARS‐CoV‐2. Innovation. 2022;3(2):100206. doi:10. 1016/j.xinn.2022.100206 Kannan SR, Spratt AN, Sharma K, Chand HS, Byrareddy SN, Singh K. Omicron SARS‐CoV‐2 variant: unique features and their impact on pre‐existing antibodies. J Autoimmun. 2022;126:102779. doi:10.1016/j.jaut.2021.102779 Saxena SK, Kumar S, Ansari S, et al. Characterization of the novel SARS‐CoV‐2 Omicron (B.1.1.529) variant of concern and its global perspective. J Med Virol. 2022 Apr;94(4):1738‐1744. doi:10.1002/ jmv.27524 Khandia R, Singhal S, Alqahtani T, et al. Emergence of SARS‐CoV‐2 Omicron (B.1.1.529) variant, salient features, high global health concerns and strategies to counter it amid ongoing COVID‐19 pandemic. Environ Res. 2022;209:112816. doi:10.1016/j.envres. 2022.112816 Montagutelli X, van der Werf S, Rey FA, Simon‐Loriere E. SARS‐ CoV‐2 Omicron emergence urges for reinforced One‐Health surveillance. EMBO Mol Med. 2022;14(3):e15558. doi:10.15252/ emmm.202115558. SUPP ORTING INFO RM ATION Additional supporting information can be found online in the Supporting Information section at the end of this article. How to cite this article: Thakur N, Das S, Kumar S, et al. Tracing the origin of severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2): a systematic review and narrative synthesis. J Med Virol. 2022;1‐14. doi:10.1002/jmv.28060