aged animals (Fig. 1 and Supplementary Fig. 3). Using low-dose infection, SHC014-MA15 failed to protect aged animals from lethal SARS-CoV challenge (Supplementary Fig. 6e, f). Together, the data suggest that SHC014-MA15 challenge can confer cross-protection against SARS-CoV through conserved epitopes, but requires a dose that induces pathogenesis. Having established SHC014 spike as a potential threat, we next synthesized a full-length SHC014-CoV infectious clone based on the approach used for SARS-CoV (Fig. 3a)15 . Replication in Vero cells revealed no deficit for SHC014-CoV relative to SARS-CoV (Fig. 3b); however, SHC014-CoV was significantly (p < 0.01) attenuated in primary human airway epithelial cultures at both 24 and 48 hours post infection (Fig. 3c). In vivo infection demonstrated no significant weight loss, but defined reduced viral replication for full length SHC014-CoV infection compared to SARS-CoV Urbani (Fig. 3d, e). Together, the results establish the viability of full length SHC014-CoV, but suggest further adaptation is required to be equivalent to epidemic SARS-CoV replication in human respiratory cells and in mice. During the SARS-CoV epidemic, links were quickly established between palm civets and coronavirus strains detected in humans2 . Building upon this finding, the common emergence paradigm argued that epidemic SARS-CoV originated as a bat virus, jumped to civets, and incorporated changes within the RBD to improve binding to civet Ace216. Subsequent exposure to humans in live markets permitted infection with the civet strain, which, in turn, adapted to become the epidemic strain (Fig. 4a). However, phylogenetic analysis suggested that early human SARS strains appear more closely related to bat than civet strains16 . Therefore, a second paradigm argued that direct bat-human transmission initiated SARSCoV emergence, with palm civets serving as a secondary host and reservoir for continued infection (Fig. 4b,17). For both paradigms, spike adaptation in a secondary host is seen as a necessity, with most mutations expected within the RBD and facilitating improved infection. Both theories imply that pools of bat CoVs are limited and host range mutations are both random and rare, reducing the likelihood of future emergence events in humans. While not invalidating the other emergence routes, the current study argues for a third paradigm in which circulating bat CoV pools maintain “poised” spike proteins capable of infecting humans without mutation or adaptation (Fig. 4c). Illustrated with SHC014 spike in Menachery et al. Page 4 Nat Med. Author manuscript; available in PMC 2016 June 01. Author Manuscript Author Manuscript Author Manuscript Author Manuscript the SARS-CoV backbone, robust infection occurs in both human airway cultures and in vivo without RBD adaptation. Coupled with previous identification of pathogenic CoV backbones1,18, the results suggest that the starting materials required for SARS-like emergent strains are currently circulating in animal reservoirs. Importantly, while full-length SHC014-CoV likely requires additional backbone adaption to mediate human disease, the documented high frequency recombination events in CoV families underscores the possibility of future emergence and the need for further preparation. To date, genomics screens of animal populations have primarily been used to identify novel viruses in outbreak settings19. The approach in this manuscript extends these datasets to examine questions of emergence and therapeutic efficacy. For the SHC014 spike, we define a threat due to replication in primary human airway cultures, the best available model for human disease. In addition, pathogenesis in mice indicates a capacity to cause disease in mammalian models without RBD adaptation. Notably, differential tropism in the lung and attenuation of full-length SHC014-CoV in HAE cultures suggest factors beyond ACE2 binding may contribute to emergence including spike processivity, receptor bio-availability, or antagonism of the host immune responses. However, further testing in non-human primates is required to translate these finding into pathogenic potential in humans. Importantly, the failure of available therapeutics defines a critical need for further study and treatment development. With this knowledge, surveillance programs, diagnostic reagents, and effective treatments can be produced to protect from emergence of group 2b specific CoVs like SHC014 as well as other CoV branches that maintain similar heterogeneous pools. While offering preparation against future emerging viruses, this approach must be considered in the context of the US government-mandated pause on gain of function (GOF) studies20. Based on previous models of emergence (Fig. 4a, b), the creation of chimeric viruses like SHC014-MA15 was not expected to increase pathogenicity. However, while SHC014-MA15 is attenuated relative to parental mouse adapted, equivalent studies examining the wild-type Urbani spike within the MA15 backbone produced no weight loss and replication attenuation21. As such, relative to the Urbani Spike-MA15 CoV, SHC014- MA15 constitutes a gain in pathogenesis (Fig. 1). Based on these findings, review panels may deem similar studies too risky to pursue as increased pathogenicity in mammalian models cannot be excluded. Coupled with