The stakes of this kind of work are hard to overstate. In the past two decades, three zoonotic coronaviruses—SARS-CoV-1, MERS-CoV, and SARS-CoV-2—have caused major outbreaks, and each emergence began with a virus crossing a species barrier. For sarbecoviruses, that barrier is defined largely by compatibility between the viral receptor-binding domain (RBD) on the spike protein and the host’s angiotensin-converting enzyme 2 (ACE2) receptor. If a virus’s RBD can engage a species’ ACE2 efficiently, that species becomes a plausible host; if it cannot, the chain of transmission breaks at the very first step. Understanding which receptors are vulnerable to which viruses is therefore central to anticipating where the next spillover might come from.

To build the atlas, the researchers first established an evolutionary framework. Phylogenetic analysis of the RBD sequences divided the 46 viruses—31 from bats, 10 from humans, three from civets, and two from pangolins—into three major clades, with clade 1 further splitting into subclades 1a and 1b. Clade 1a contains SARS-CoV-1 and related bat and civet strains; clade 1b contains SARS-CoV-2 and its closest relatives, including the bat virus RaTG13 and pangolin coronaviruses. Clade 2, which comprises most known sarbecoviruses, carries a distinctive double-segment deletion in the receptor-binding motif, while clade 3 viruses from African and European bats carry a single-segment deletion in the same region. These structural differences in the RBM, the loop that physically contacts ACE2, turned out to predict much of the functional variation the team later observed.

The functional screen itself relied on pseudotyped reporter viruses: vesicular stomatitis virus particles decorated with each sarbecovirus spike protein, whose entry into cells expressing a given ACE2 ortholog could be quantified by luminescence. The team confirmed comparable spike expression and incorporation across pseudovirus preparations by western blot and normalized viral input by quantitative RT-PCR, allowing a fair comparison across the roughly 3,000 possible virus-receptor pairings. The result is a heatmap of cross-species infectivity that functions as a kind of vulnerability index for each species’ receptor.

Clade 1 viruses emerged as the clear standouts in receptor breadth. Within clade 1b, the pangolin viruses PCoV-GD and PCoV-GX, which share 89.6 and 92.3 percent spike identity with SARS-CoV-2 respectively, showed broad ACE2 compatibility, with PCoV-GD efficiently infecting nearly all tested non-bat mammalian ACE2s and most bat ACE2s. Intriguingly, RaTG13—despite sharing 97.4 percent identity with SARS-CoV-2 across the full spike protein—displayed a more restricted host range, implying that subtle differences within the receptor-binding motif underlie the expanded tropism of SARS-CoV-2. Within clade 1a, human SARS-CoV-1 isolates used ACE2 orthologs from a wide range of bat families, whereas most bat- and civet-derived relatives showed narrower usage, with the bat virus WIV1 bridging human and bat lineages through its ability to engage ACE2 from multiple Rhinolophus and Myotis species.

Clade 2 and clade 3 viruses told very different stories. None of the clade 2 pseudoviruses, which represent the majority of known sarbecoviruses, mediated entry through any of the 66 ACE2 orthologs, consistent with their double RBM deletion and previously reported ACE2-independent entry mechanisms. Clade 3 viruses showed intermediate phenotypes: BtKY72 used ACE2 from only a few Rhinolophus species and did not engage human ACE2, while Khosta-2 showed broader tropism, infecting cells expressing rabbit ACE2 more than ten-fold more efficiently than those expressing the human receptor—a striking example of lineage-specific adaptation.

The study also delivered a decisive verdict on several proposed alternative receptors. Host factors including NRP1, CD147, AXL, and TMEM106B have each been suggested to facilitate SARS-CoV-2 entry in various experimental contexts. Yet across the entire 46-virus panel, none of these factors supported detectable infection when overexpressed in the assay cells, indicating that none functions as an independent entry receptor for any sarbecovirus tested. The authors are careful to note that context-dependent accessory roles in specific SARS-CoV-2 settings cannot be excluded, but the message is clear: ACE2 remains the only demonstrated primary entry receptor across this viral diversity.

Among the most consequential findings is the identification of two bat species whose receptors behave remarkably like the human version. Only the ACE2 orthologs of Rhinolophus affinis and Miniopterus schreibersii correlated strongly with human ACE2 usage across the viral panel, with correlation coefficients of 0.8314 and 0.8777 respectively. Surface plasmon resonance confirmed that M. schreibersii ACE2 binds the SARS-CoV-1 and SARS-CoV-2 RBDs with high affinity—dissociation constants of 8.14 × 10⁻⁸ M and 8.06 × 10⁻⁷ M, lower than human ACE2’s nanomolar binding but fully consistent with efficient entry. Because both species range across Southeast Asia, Europe, and North Africa in regions overlapping human habitats, the authors argue they should be priorities for ongoing surveillance. The data also flagged rabbits, cattle, sheep, raccoon dogs, badgers, Siberian chipmunks, and pigs as mammalian species whose ACE2 usage correlates strongly with human ACE2, marking them as candidate intermediate hosts worth investigating.

Perhaps the most mechanistically revealing part of the study concerns two closely related Miniopterus bats whose receptors sit at opposite ends of the permissiveness spectrum. M. schreibersii ACE2 supported entry by nearly all ACE2-dependent pseudoviruses, while the ACE2 of M. natalensis was entirely non-permissive—despite the two proteins differing by only seven amino acids. Three of those substitutions, at positions 27, 31, and 42, sit directly at the predicted RBD-binding interface. Molecular dynamics simulations using the molecular mechanics generalized Born surface area method showed that mutating M. schreibersii ACE2 residue I27 to lysine raised the binding free energy by roughly 5 kcal/mol, while the K42E substitution raised it by nearly 7 kcal/mol, disrupting a salt-bridge network involving residue D38 and the SARS-CoV-1 RBD residue Y442. Entry assays confirmed the simulations: the I27K mutation selectively impaired civet-derived SARS-CoV-1 entry, N31G had minimal impact, and K42E broadly abolished usage by nearly all pseudotypes. Reciprocally, introducing the corresponding M. schreibersii residues into M. natalensis ACE2 restored entry competence for multiple SARS-CoV-1 pseudotypes. Residues 27 and 42, in short, are the principal molecular switches governing Miniopterus receptor recognition.

The authors are candid about the limits of their approach. Pseudoviruses built from available sequences may not capture the full diversity of natural sarbecoviruses, and receptor compatibility is only one barrier to cross-species transmission; protease availability, replication competence, and immune evasion all matter for productive infection in a living animal. Efficient ACE2 engagement alone does not guarantee disease. Still, the atlas provides exactly the kind of mechanistic framework that field surveillance has lacked: a way to prioritize which of the world’s more than 1,400 bat species, and which of the hundreds of circulating sarbecovirus sequences, warrant the closest monitoring. As the authors put it, identifying species with ACE2 structures resembling the human receptor may help focus receptor-based risk assessment where it matters most—before, rather than after, the next spillover.

Subject of Research: Cross-species ACE2 receptor usage and host-range determinants of sarbecoviruses

Article Title: An atlas of sarbecovirus infectivity across diverse ACE2 orthologs

Article References: Sun, Y., Cheng, Z., Wu, X., Liu, K., Wang, L., Yang, D., Huang, W., & Nie, J. (2026). An atlas of sarbecovirus infectivity across diverse ACE2 orthologs. iScience, 29(10), Article 117769. https://doi.org/10.1016/j.isci.2026.117769

Image Credits: AI Generated

DOI: 10.1016/j.isci.2026.117769

Keywords: sarbecovirus, ACE2, SARS-CoV-2, SARS-CoV-1, bats, spillover, pseudovirus, receptor-binding domain, host range, surveillance, Miniopterus, Rhinolophus