It has been two months since the outbreak of the coronavirus in Wuhan and its spread has shown no signs of slowing down in China. More than 35 Chinese cities have been put on lockdown by Chinese authorities in an attempt to isolate confirmed and suspected cases. The lives of millions of people are in danger as the virus shows signs of spreading further in China as well as internationally.
There are significant gaps in the official investigations into the origins of the novel Coronavirus. In order to contain the virus, one first needs to understand how a virus that allegedly originated in animals found its way to humans. For this to happen, the Chinese authorities need to release their animal testing data and samples. Testing results from animal samples collected at epicenters would give important insights into what animals might serve as intermediate hosts for the new coronavirus.
An Animal Origin of the VirusScientific studies based on phylogenetic analysis have researched the sequence of the novel coronavirus, compared it to other coronavirus sequences, and found it likely originated in bats. Researchers from the Wuhan Institute of Virology found the genome in the virus found in patients was 96 percent identical to that of an existing bat coronavirus, according to a study published in the journal Nature. But there have been other theories as well. One Chinese study suggested, for example, that snakes were the source of transmission to humans. However, many scientists believe that reptiles are a less likely source and that mammals like rats and pigs, and some birds, have been the primary reservoir for coronaviruses.
With this in mind, phylogenetic studies of viral genome sequences need to be supported by animal studies to confirm the origin of the infection, as well as to determine whether there is an intermediate host.
It is not an easy task for a virus to establish zoonotic transmission, and coronaviruses rarely leap from animal to human infection with high transmissibility. There is even less chance to see a coronavirus leap directly from bats to humans. To infect new hosts, mutations need to occur with the viral surface proteins and/or envelope and structural genes, so that the mutated viruses can bind and enter the cells of new species, and efficiently complete the replication cycles in the new hosts.
Some scientists have argued that coronaviruses can jump directly to humans, without mutating or passing through an intermediate species. However, an intermediate host was clearly needed to establish zoonotic transmission to humans in the previous outbreaks of coronaviruses. Many studies suggested that the bat coronavirus jumped from its natural host bats to civets and then to humans during the 2003 SARS outbreak, and it jumped from bats to camels and then to humans for the MERS outbreak. So, civets and camels would serve as intermediate hosts for zoonotic transmission.
Because bats were not sold at the Huanan market in Wuhan—the epicenter of the infection—at the time of the outbreak, this suggests the existence of another intermediate animal host that may have transferred the virus to humans.
What is the most puzzling is that there have been no reports on the testing of animal samples collected in any epicenters in Wuhan, especially at the Huanan seafood market, to identify what animals might be the host or intermediate hosts of this novel Wuhan coronavirus.
Chinese scientists published a report in Lancet recently which stated that “the majority of the earliest cases included reported exposure to the Huanan Seafood Wholesale Market” and that patients could have been infected through zoonotic or environmental exposures. Another report on Lancet by Chinese CDC scientists claimed that “on the basis of current data, it seems likely that the 2019-nCoV causing the Wuhan outbreak might also be initially hosted by bats, and might have been transmitted to humans via currently unknown wild animal(s) sold at the Huanan seafood market.”
However, so far, no information was released about the amount, and species, of wild animals present at the Huanan seafood market upon closure; nor about how the animals were managed or disposed of when the market was closed on Jan. 1, 2020. And no information was released about how many animal samples were tested for SARS-CoV or Wuhan Coronavirus via viral nucleic acid testing methods.
Official Chinese state news agency Xinhua reported on Jan. 26 that 33 samples out of 585 environmental samples collected at the Huanan Seafood market were positive for nucleic acids from new Coronavirus, suggesting the virus originated from wild animals or stocks sold there. However, these samples were from the environment—not from animals.
Background of the Huanan Seafood Market ClosureThe 2019-nCoV has caused rapid infection in China and spread to other countries outside China, which has led to a global health crisis.
No Information on Wild Animals at the Seafood Market Was DisclosedYet, no information was released about the amount, and species, of animals present upon closure, how many animals were tested for Coronavirus, and how the animals were managed or disposed of upon the closure of the market on Jan. 1. A Chinese media outlet, Yicai, inquired about the outcome of the wild animals sold at the market and confirmed that there was no disclosure from the Wuhan government.
The Huge Risks of Not Identifying the Original or Intermediate Animal HostsThe U.S. Center for Disease Control (CDC) stated that “much is unknown about how 2019-nCoV, a new coronavirus, spreads.” So far the understanding is that the major pathway of 2019-nCoV infection is respiratory droplet transmission and contact from humans to humans.
Guan Yi and Kwok-yung Yuen of the University of Hong Kong (HKU) et. al. identified severe acute respiratory syndrome coronavirus (SARS-CoV) from caged palm civets from live animal markets in China in 2003. Their studies lead to the subsequent ban on selling civets and the closing of all wild animal markets in Guangdong and helped to confine the SARS epidemic.
Huanan seafood market is also infested by rodents. If rodents were elucidated as being a potential host for coronavirus, the risk of rats infesting beyond the current quarantine zone still persists. Given the fact that coronavirus was detected from feces from patients from Shenzhen and that bat SARS-like virus strains were isolated from bat feces, the possible fecal-oral route of 2019-nCoV transmission in addition to respiratory droplet transmission would lead to a reasonable warning for people to avoid contact with animals like rats. Thus, if rodents are indeed a source or host of the 2019-NCoV infection, then, rodent contamination of food or water is a potential way for the disease to spread, which needs to be brought to the awareness of the international community.
Similarly, if birds or other species were the hosts of 2019-nCoV in the seafood market, the information pertaining to the species, amount, virus type, biological reactions, and potential routes of spreading of the virus also need to be identified or reported to the world so that appropriate prevention measures could be taken.
Were There Other Epicenters Besides the Huanan Seafood Market?The Chinese CDC did release data from environmental samples from the seafood market and suggested that “it is originated from wild animals with species uncertain.”
The Possibility 2019-nCoV Originated From Bat SARS-Like Virus (Bat-SL-CoV)One recent Lancet report on Jan. 29, 2020, titled "Genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding," stated that “A Blast search of the complete genomes of 2019-nCoV revealed that the most closely related viruses available on GenBank were bat-SL-CoV-ZC45 (sequence identity 87.99%; query coverage 99%) and another SARS-like betacoronavirus of bat origin, bat-SL-CoV-ZXC21 (accession number MG772934;23 87.23%;” “Notably, the 2019-nCoV strains were less genetically similar to SARS-CoV (about 79%) and MERS-CoV (about 50%).”
This message might be interpreted as 2019-nCoV being biologically closer related to SARS-like betacoronavirus of Bat origin and bats may be the original host of this virus. However, the authors did not claim that the only host to 2019-nCoV is a bat.
The paper stated that “However, despite the importance of bats, several facts suggest that another animal is acting as an intermediate host between bats and humans. First, the outbreak was first reported in late December, 2019, when most bat species in Wuhan are hibernating. Second, no bats were sold or found at the Huanan seafood market, whereas various non-aquatic animals (including mammals) were available for purchase. Third, the sequence identity between 2019-nCoV and its close relatives bat-SL-CoVZC45and bat-SL-CoVZXC21 was less than 90%, which is reflected in the relatively long branch between them. Hence, bat-SL-CoVZC45 and bat-SL-CoVZXC21 are not direct ancestors of 2019-nCoV. Fourth, in both SARS-CoV and MERS-CoV, bats acted as the natural reservoir, with another animal (masked palm civet for SARS-CoV35 and dromedary camels for MERS-CoV) acting as an intermediate host, with humans as terminal hosts. Therefore, on the basis of current data, it seems likely that the 2019-nCoV causing the Wuhan outbreak might also be initially hosted by bats, and might have been transmitted to humans via currently unknown wild animal(s) sold at the Huanan seafood market.”
Studies From Wuhan Institute of Virology on Bat SARS-Like CoV.Zheng-Li Shi and several other researchers at the Wuhan Institute of Virology published an article in Nature in 2013 titled "Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor.”
In that study, their team harvested from anal swabs or fecal samples from bats and found 2 strains of sequences from Bat SARS-Like CoV that termed as RsSHC014 and Rs3367. They process 95% nucleotide sequence identity with human SARS-CoV Tor2 strain.
They collected serum from 218 residents who live close to bat caves with bats carrying the viruses. Those caves were the places where the Shi group collected the virus samples. Then ELISA assays were conducted to detect antibodies to bat SARS-CoV, since antibody existence would suggest a prior exposure to the bat coronavirus. They found that only 6 out of 218 (2.7 %) subjects showed seropositivity, which suggested likely infections to bat SARS-CoVs or related viruses. No clinical symptoms have been manifested in the 6 positive persons in the past 12 months. As a control, they collected 240 samples from random blood donors in Wuhan, 1000 km away from Yunnan, none of the Wuhan blood samples showed any positivity to bat SARS-like CoV.
Track Record of Wuhan Institute of Virology on Engineering ‘Gain-of-Function’ Bat SARS-Like CoV.Zhengli Shi’s group at the Institute of Virology at Wuhan was successful in isolating two infectious clones of bat SARS-Like CoV: SL-CoV-WIV1 and WIV16 from bats. In their further studies, they found out that these SL-CoV Spike protein (S protein) “[were] unable to use any of the three ACE2 molecules as its receptor; Second, the SL-CoV failed to enter cells expressing the bat ACE2; Third, the chimeric S covering the previously defined receptor-binding domain gained its ability to enter cells via human ACE2, albeit with different efficiencies for different constructs; Fourth, a minimal insert region ( Amino acids 310 to 518 ) was found to be sufficient to convert the SL-CoV S from non-ACE2 binding to human ACE2 binding.”
Furthermore, Shi’s group joined an international group to generate a chimeric virus with the bat virus SHC014 they harvested in Yunnan. Since they know SHC014 is unlikely to bind to human ACE2, they “synthesized the SHC014 spike in the context of the replication competent, mouse-adapted SARS-CoV backbone”. So, that is a lab-engineered virus with SARS-CoV Mouse adapted backbone (MA15) but with SHC014 spike.
To their surprise, the chimeric virus (SHC014-MA15) can use SHC014 spike to bind to human ACE2 receptor and enter human cells. SHC014-MA15 can also cause disease in mice and cause death as well. Existing vaccines to SARS cannot protect animals from SHC014-MA15 infection. Therefore, these chimeric virus studies can lead to the generation of more pathogenic, more deadly CoV strains in mammalian models.
Due to the U.S. government-mandated pause on the gain-of-function (GOF) studies, this international research did not proceed further at that time. However, there is no evidence that Shi’s group in China stopped any further study on the track of introducing GOF mutations on the CoV. And it is clear that Shi’s group already mastered the reverse-engineering technology that is sufficient to introduce mutation in current SARS-CoV or SARS-Like CoV to create mutant infectious coronavirus.
Possibilities of an Animal Host of 2019-nCoV at Huanan Seafood MarketOne recent Lancet paper titled "Genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding," reported that “as a typical RNA virus, the average evolutionary rate for coronaviruses is roughly 10⁴ nucleotide substitutions per site per year, with mutations arising during every replication cycle. It is, therefore, striking that the sequences of 2019-nCoV from different patients described here were almost identical, with greater than 99.9% sequence identity. This finding suggests that 2019-nCoV originated from one source within a very short period and was detected relatively rapidly.”
With mutations in every cycle, it is highly unlikely for different bats to host viruses with the same sequence. If bats alone are not enough for virus transmission, another animal is needed as the intermediate host, and the chance of the virus being identical is even slimmer. Since the seafood market is not the only source for the outbreak, it is reasonable to postulate that if another animal is the intermediate host for the virus, that animal needs to have contact with bats, allow bat coronavirus to proliferate in them, and, eventually, the animal needs to have the capacity to transmit viruses to human beings who may or may not have contact with the Huanan seafood market.
Therefore, there have been serious questions on whether this Wuhan coronavirus outbreak was due to a leak or mishandling of laboratory animals used in coronavirus studies. This is a reasonable public inquiry regarding the source of the outbreak and it warrants a transparent investigation from the Chinese authorities and foreign disease control and laboratory operation experts. This is not just about the accountability of medical ethics or laboratory safety operations, it is directly related to the current endeavors to contain the virus outbreak.
While the animal host of 2019-nCoV is yet to be identified, the data and information from possible animal hosts and potential zoonic infection is imperative for prevention and controlling disease on an international scale.
The Huanan seafood market has a high potential of harboring the animal host. Animal data and profiling results from the Huanan seafood market need to be disclosed immediately by Chinese authorities even if they are negative results. It is imperative for U.S. CDC and WHO officers to demand that Chinese authorities release the information about animal testing data.
If Chinese authorities refuse to disclose testing data for animal samples, it could imply an intentional cover-up of the true origin of the 2019-nCoV outbreak.