Edited by: Yashpal S. Malik, Indian Veterinary Research Institute (IVRI), India
Reviewed by: Santhamani Ramasamy, Albert Einstein College of Medicine, United States; Tung Phan, University of Pittsburgh Medical Center, United States; Sharvan Sehrawat, Indian Institute of Science Education and Research Mohali, India
This article was submitted to Virology, a section of the journal Frontiers in Microbiology
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The coronavirus disease 19 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic since the first report in Wuhan. COVID-19 is a zoonotic disease and the natural reservoir of SARS-CoV-2 seems to be bats. However, the intermediate host explaining the transmission and evolvement is still unclear. In addition to the wildlife which has access to contact with bats in the natural ecological environment and then infects humans in wildlife market, domestic animals are also able to establish themselves as the intermediate host after infected by SARS-CoV-2. Although recent studies related to SARS-CoV-2 have made a lot of progress, many critical issues are still unaddressed. Here, we reviewed findings regarding the investigations of the intermediate host, which may inspire future investigators and provide them with plenty of information. The results demonstrate the critical role of the intermediate host in the transmission chain of SARS-CoV-2, and the efficient intervention on this basis may be useful to prevent further deterioration of COVID-19.
The current global pandemic of coronavirus disease 19 (COVID-19) is attributed to the transmission of a pathogen named SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2). In the early stage of the outbreak of COVID-19,
The potential transmission of SARS-CoV-2 between hosts and humans. SARS-CoV-2, originated from bat-nCoV, infected wild animals and gradually evolved in the intermediate host after mutation and recombination. Wildlife business give chance for SARS-CoV-2 to infect humans and domestic animals. SARS-CoV-2 induce a pandemic in human population by respiratory droplet transmission and close contact transmission.
Currently, SARS-CoV-2 is a member of the genus Betacoronavirus family, of which the severe acute respiratory syndrome coronavirus (SARS-CoV) and the Middle East respiratory syndrome coronavirus (MERS-CoV) have caused epidemics in humans before (
Since
The zinc finger antiviral protein (ZAP), an important mammalian antiviral protein, can bind specifically to CpG dinucleotides and degrade viral genomes (
Although Bat-nCoV RaTG13 share 96.2% similarity with SARS-CoV-2, the identity of RBD to SARS-CoV-2 is inferior to pangolin-nCoV. The analysis of pangolin-nCoV metagenomic dataset and the multiple alignments across the RBM segments revealed that pangolin-nCoV shared 89% nucleotide similarity and 98% amino acid identity to SARS-CoV-2 (
The S protein played a crucial role in determining host tropism and transmission capacity. It consists of S1 domain mediating receptor binding and S2 domain responsible for cell membrane fusion. With the help of cellular protease such as transmembrane protease serine2 (TMPRSS2), efficient proteolytic processing of S protein at the S1/S2 cleavage site is also important in mediating SARS-CoV-2 entry, after which S2 subunit will allow fusion of viral and cellular membranes (
On such a premise that virus acquires the capability of efficient replication by evolving similar codon usage pattern to its hosts, the high similarity of the relative synonymous codon usage (RSCU) bias between SARS-CoV-2 and snakes found by
According to current understanding, ferrets are susceptible to infect SARS-CoV, whereas at the same time avoid the MERS-CoV invasion (
Deng et al. performed an antibody survey among 1914 serum samples from 35 species and did not detect any SARS-CoV-2 specific antibody. They consequently draw a conclusion that those species can be excluded from the candidates for the intermediate host (
Based on the view that key atomic-level interaction for coronavirus invasion is between the RBD and host ACE2 receptor, lots of studies were performed surrounding key residues (
Functional assays of ACE2 protein using authentic SARS-CoV-2 conducted by
Currently, the bat is the natural host for SARS-CoV-2 according to mainstream perspectives, but the intermediate host of SARS-CoV-2 is still unclear (
The potential intermediate hosts for SARS-CoV-2.
Pangolin | Gene Sequence Analysis and Comparison | Malayan pangolins contain sequences strongly similar to SARS-CoV-2 | |
High-throughput Sequencing and phylogenetic analysis | Pangolin-nCoV belongs to two sub-lineages of SARS-CoV-2 related coronaviruses | ||
Molecular and phylogenetic analyze the assembled complete genome of pangolin-nCoV | Pangolin-nCoV have the highly conserved S genes and structure of RBD protein to SARS-CoV-2 | ||
Gene sequence analysis of coronavirus genomes reconstructed from viral metagenomic datasets of hosts and SARS-CoV-2 | High sequence similarity in the RBM between SARS-CoV-2 and a coronavirus genome from datasets of pangolin | ||
Systematic comparison and analysis to predict the interaction between the RBD and the ACE2 | Regarding the similarity of the key amino acids of interaction between RBD and ACE2 to humans, the pangolin is closer than the bat | ||
Molecular evolution and phylogenetic analysis of SARS-CoV-2 and hosts ACE2 protein | The evolutionary divergence between pangolin ACE2 and hACE2 is lower than that between bat ACE2 and hACE2 | ||
The phylogenetic tree based on Hausdorff distance and Center distance between SARS-CoV-2 strains and host-nCoV groups | The pangolin-nCoV is closely related to the SARS-CoV-2 group based on the genome divergences | ||
Phylogenetic, split network, transmission network, and comparative analyses of the genomes | The pangolin-nCoV from the two pangolin samples did not have the PRRA insertion, which is crucial in viral invasion | ||
Virus infectivity studies using HEK293T cells expressing ACE2 from 11 species of animals | Pangolin ACE2 could mediate SARS-CoV-2 entry | ||
Pseudotyping particles of Spike mimics particle entry and quantitative cell-cell fusion assay | Pangolin sustained higher levels of entry than was seen with an equivalent hACE2 construct | ||
Mink | Compare the infectivity patterns by deep learning algorithm of VHP | Mink coronavirus have the closest infectious patterns to SARS-CoV-2 | |
Genetic and epidemiological sleuthing | The SARS-CoV-2 outbreak in mink farms is introduced by humans, and infected minks can transmit the virus to human and other animals via viral dust or droplets | ||
Turtle | Systematic comparison and analysis to predict the interaction between the RBD and ACE2 | Regarding the similarity of the key amino acids of interaction between RBD and ACE2 to humans, the turtle is closer than the bat | |
Analyze the affinity to S protein of the 20 key residues in ACE2 | |||
Snake | Relative synonymous codon usage (RSCU) comparison and analysis | Snake shared the lowest RSCU distance to SARS-CoV-2 | |
Analyze the affinity to S protein of the 20 key residues in ACE2 | |||
Reperform the RSCU comparison and analysis conducted by |
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Ferrets | Establish a ferret model of SARS-CoV-2 infection and transmission | SARS-CoV-2 is effectively transmitted to naïve ferrets by direct contact and leads acute bronchiolitis | |
Intranasally inoculated SARS-CoV-2 to domestic animals | Ferrets have high susceptibility to SARS-CoV-2 | ||
Pseudotyping particles of Spike mimics particle entry and quantitative cell-cell fusion assay | Ferret ACE2 is not used efficiently by SARS-CoV-2 for entry | ||
Bovidae (yak) | Analyze the affinity to S protein of the 20 key residues in ACE2 | The majority of key residues in ACE2 are identical to hACE2 protein | |
Phylogenetic tree analysis and structural models’ comparison | A yak betacoronavirus strain has spike glycoproteins structure models closest to SARS-CoV-2 | ||
Dogs | Pseudotyping particles of Spike mimics particle entry and quantitative cell-cell fusion assay | Dog sustained higher levels of entry than was seen with an equivalent hACE2 construct | |
Intranasally inoculated SARS-CoV-2 to domestic animals | |||
Investigate the level of ACE2 expression in different organs | |||
Use single-cell technique to screen of ACE2 and TMPRSS2(SARS-CoV-2 target cell) in different organs of animals | |||
Virus infectivity studies using HEK293T cells expressing ACE2 from 11 species of animals | Dog ACE2 could mediate SARS-CoV-2 entry | ||
Cats | Intranasally inoculated SARS-CoV-2 to domestic animals | Cats have high susceptibility to SARS-CoV-2 | |
Phylogenetic clustering and sequence alignment to evaluate the receptor-utilizing capability of ACE2 | Cat ACE2 have the receptor-utilizing capability of SARS-CoV-2 | ||
Use expressed RBD proteins to perform surface staining of cells transfected with expression plasmids of ACE2 orthologs | Cats support the efficient entry of SARS-CoV-2, SARS-CoV, and Bat-nCoV RaTG13 | ||
Intranasally inoculated SARS-CoV-2 or close contact with infected cat | Cats are subclinical infection and shed virus for no more than 5 days | ||
Pseudotyping particles of Spike mimics particle entry and quantitative cell–cell fusion assay | Cat sustained higher levels of entry than was seen with an equivalent hACE2 construct | ||
Use single-cell technique to screen of ACE2 and TMPRSS2(SARS-CoV-2 target cell) in different organs of animals | Cats have the highest proportion of SARS-CoV-2 target cell, and those cells were widely distributed among digestive system, respiratory system and urinatory system | ||
Virus infectivity studies using HEK293T cells expressing ACE2 from 11 species of animals | Cat ACE2 could mediate SARS-CoV-2 entry | ||
Swine (pigs) | Phylogenetic clustering and sequence alignment to evaluate the receptor-utilizing capability of ACE2 | Swine ACE2 have the receptor-utilizing capability of SARS-CoV-2 | |
Use expressed RBD proteins to perform surface staining of cells transfected with expression plasmids of ACE2 orthologs | Pigs support the efficient entry of SARS-CoV-2, SARS-CoV, and Bat-nCoV RaTG13 | ||
Use single-cell technique to screen of ACE2 and TMPRSS2 (SARS-CoV-2 target cell) in different organs of animals | Pig have a variety of cell types co-expressing SARS-ACE2 and TMPRSS2 | ||
Virus infectivity studies using HEK293T cells expressing ACE2 from 11 species of animals | Pig ACE2 could mediate SARS-CoV-2 entry | ||
Intranasally inoculated SARS-CoV-2 to domestic animals | |||
Investigate the level of ACE2 expression in different organs |
B-pT contributed the conception and design of this review. JZ wrote the manuscript. B-pT and WC revised and edited this manuscript. All authors reviewed the draft and approved the submission.
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
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