Functional and Genetic Analysis of Viral Receptor ACE2 Orthologs Reveals a Broad Potential Host Range of SARS-CoV-2

Abstract

The pandemic of a newly emerging coronavirus (SARS-CoV-2), the causative agent of severe pneumonia disease , is a major global health threat. Epidemiological studies suggest that bats are the natural zoonotic reservoir for SARS-CoV-2, however, the host range of SARS-CoV-2 and the identity of intermediate hosts that may facilitate the transmission to humans remains unknown. Coronavirus-receptor interaction is a key genetic determinant of the host range, cross-species transmission, and tissue tropism. SARS-CoV-2 uses Angiotensin-converting enzyme II (ACE2) as the receptor to enter its host cells in a species-dependent manner. It has been shown that human, palm civet, pig and bat ACE2 can support virus entry, while the murine ortholog cannot. In this study, we aimed to characterize ACE2 from diverse species for its ability to support viral entry. We found that ACE2 is expressed in a wide range of host species, with high conservation especially in mammals. By analyzing critical amino acid residues in ACE2 for virus entry, based on the well-characterized SARS-CoV spike protein interaction with ACE2 (human, bat, palm civet, pig and ferret ACE2), we identified approximately eighty ACE2 proteins from mammals could potentially function as the receptor to mediate SARS-CoV-2 entry. Functional assays showed that 44 of these mammalian ACE2 orthologs, including domestic animals, pet animals, livestock animals and even animals in the zoos or aquaria, could bind viral spike protein and support SARS-CoV-2 entry. In summary, our study demonstrates that ACE2 from a remarkably broad range of species support SARS-CoV-2 entry. These findings highlight a potentially broad host tropism and suggest that SARS-CoV-2 might be distributed much more widely than previously recognized, emphasizing the necessity to monitor the susceptible hosts, especially their potential of cross-species, which could prevent the future outbreaks.

SARS-CoV-2 spike protein predicted to form complexes with host receptor protein orthologues from a broad range of mammals

“…As shown in previous studies, and supported by experimental data, many primates are predicted to be at high risk [19,41,42]. In agricultural settings, camels, cows, sheep, goats and horses also have relatively low ΔΔG values, suggesting comparable binding affinities to humans, in agreement with experimental data [20,21]. In domestic settings, dogs [18], cats [18], hamsters [20], and rabbits [20][21][22] also have ΔΔG values suggesting risk, again in agreement with experimental data (Table 1).…”

“…In agricultural settings, camels, cows, sheep, goats and horses also have relatively low ΔΔG values, suggesting comparable binding affinities to humans, in agreement with experimental data [20,21]. In domestic settings, dogs [18], cats [18], hamsters [20], and rabbits [20][21][22] also have ΔΔG values suggesting risk, again in agreement with experimental data (Table 1). Whilst, zoological animals that come into contact with humans, such as pandas, leopards and bears, are also at risk of infection as shown experimentally [20] In predicting susceptibility, we have chosen thresholds supported by in vivo or in vitro experimental data.…”

“…Two animals are outliers in the infected boxplot, corresponding to horseshoe bat (ΔΔG = 3.723) and marmoset (ΔΔG = 3.438). To be cautious, since in vivo experiments have shown that marmosets can be infected, and in vitro experiments have shown that horseshoe bats can be infected [2,[20][21][22] (Table 1), we consider animals that have ΔΔG values less than, or equal to, the ΔΔG = 3.7 for horseshoe bat to be at risk. Additionally, there is a clear sampling bias in the set of animals that have so far been experimentally characterised: all but chicken and duck are mammals.…”

“…Animal infection studies have also identified cats [18] and dogs [18] as hosts, as well as ferrets [18], macaques [19] and marmosets [19]. Recent in vitro studies have also suggested an even broader set of animals may be infected [20][21][22]. To understand the potential host range of SARS-CoV-2, the plausible extent of zoonotic and anthroponotic transmission, and to guide surveillance efforts, it is vital to know which species are susceptible to SARS-CoV-2 infection.…”

“…For example, variations in other proteins may prevent downstream events that are required for viral replication in a new host. Hence, examples of real-world infections [14][15][16][17] and experimental data from animal infection studies [18][19][20][21][22] are required to validate hosts that are predicted to be susceptible.…”