Fauci Intelligence Community Release — page 46

these adaptations would enable the epidemic to take off, producing a sufficiently large cluster of cases to trigger the surveillance system that detected it​1,2​. All SARS-CoV-2 genomes sequenced so far have the genomic features derived above and are thus derived from a common ancestor that had them too. The presence in pangolins of an RBD very similar to that in SARS-CoV-2 means we can infer this was also likely in the virus that jumped to humans. This leaves the polybasic cleavage site insertion to occur during human-to-human transmission. Estimates of the timing of the most recent common ancestor of SARS-CoV-2 using current sequence data point to virus emergence in late November to early December 2019​23​, compatible with the earliest retrospectively confirmed cases​24​. Hence, this scenario presumes a period of unrecognised transmission in humans between the initial zoonotic event and the acquisition of the polybasic cleavage site. Sufficient opportunity could occur if there had been many prior zoonotic events producing short chains of human-to-human transmission over an extended period. This is essentially the situation for MERS-CoV where all human cases are the result of repeated jumps of the virus from dromedary camels, producing single infections or short transmission chains that eventually resolve, with no adaptation to sustained transmission​25​. Studies of banked human samples could provide information on whether such cryptic spread has occurred. Retrospective serological studies could also be informative and a few such studies have been conducted showing low-level exposures to SARS-like coronaviruses in certain areas of China ​26​. Critically, however, these studies could not have distinguished whether exposures were due to prior infections with SARS-CoV, SARS-CoV-2, or other SARS-like coronaviruses. Further serological studies should be conducted to determine the extent of prior human exposure to SARS-CoV-2. 3. Selection during passage Basic research involving passage of bat SARS-like coronaviruses in cell culture and/or animal models have been ongoing in BSL-2 for many years in laboratories across the world​27 and there are documented instances of laboratory escapes of SARS-CoV​28​. We must therefore examine the possibility of a inadvertent laboratory release of SARS-CoV-2. In theory, it is possible that SARS-CoV-2 acquired RBD mutations (​Fig. 1a​) during adaptation to passage in cell culture, as has been observed in studies with SARS-CoV​11​. The finding of SARS-like coronaviruses from pangolins with near-identical RBDs, however, provides a much stronger and parsimonious explanation for how SARS-CoV-2 acquired these via recombination or mutation​19​. The acquisition of both the polybasic cleavage site and predicted O-linked glycans also argues against culture-based scenarios. New polybasic cleavage sites have only been observed after prolonged passage of low pathogenicity avian influenza virus ​in vitro or ​in vivo​17​. Furthermore, a hypothetical generation of SARS-CoV-2 by cell culture or animal passage would have required prior isolation of a progenitor virus with very high genetic similarity, which has not been described. Subsequent generation of a polybasic cleavage site would have then required repeated passage in cell culture or animals with ACE2 receptors similar to humans, but such work has also not previously been described. Finally, the generation of the predicted O-linked glycans is also unlikely to have occured due to cell culture passage, as such features suggest the involvement of an immune system​18​. Conclusions In the midst of the global COVID-19 public health emergency it is reasonable to wonder why the origins of the epidemic matter. A detailed understanding of how an animal virus jumped species boundaries to infect humans so productively will help in the prevention of future zoonotic events. For example, if SARS-CoV-2 pre-adapted in another animal species then we are at risk of future re-emergence events. In contrast, if the adaptive process occurred in humans, then even if we have repeated zoonotic transfers they are unlikely to take off without the same series of mutations. In addition, identifying the closest animal relatives of SARS-CoV-2 will greatly assist studies of virus function. Indeed, the availability of the RaTG13 bat sequence helped reveal key RBD mutations and the polybasic cleavage site.