Laboratory Cell Culture Footprints in the SARS-CoV-2 Glycan Shield

Laboratory Cell Culture Footprints in the SARS-CoV-2 Glycan Shield The origin of SARS-CoV-2 is still disputed. Structural and evolutionary evidence often point in different directions. Early arguments for a natural origin, especially those in Andersen et al. (2020), claimed that features such as the furin cleavage site and its predicted O-linked glycans could only have evolved under pressure from a living animal’s immune system. Later findings have weakened this claim. The predicted O-linked glycans do not form the protective shield originally described, and their presence does not rule out adaptation in cell culture. Laboratory passaging in different cell lines puts strong selective pressure on the spike protein, even without an immune system. In Vero E6 cells, which lack the TMPRSS2 enzyme, the furin cleavage site is quickly lost. In human lung and intestinal cells such as Calu-3 and Caco-2, the same site is kept and refined. At the same time, SARS-CoV-2 is missing a conserved glycan shield at position N370 because of a mutation called T372A. When researchers put this glycan back, the spike locks shut and the virus replicates more than 60 times less efficiently in human lung cells. Interestingly, the same uncommon codon for alanine (GCA) appears in both the T372A mutation and the furin cleavage site insertion. Pre-2020 laboratory work had already established methods for removing specific glycan shields, including the precise structural equivalent of the N370 site. Vaccine-oriented studies by collaborators of the Wuhan Institute of Virology deliberately ablated this glycan (mutating the 372 site to alanine), and the 2018 DEFUSE proposal explicitly planned similar N-glycan manipulations to study host-range effects. Recent evolutionary analyses (Havens et al., 2026) found no clear signs of strong natural selection on the virus’s lineage just before it appeared in humans. This suggests that SARS-COV-2 already carried these human-adapted features when it first entered the human population. There are known methodological and interpretive problems with the Havens study (Bostickson, 2026a). Even so, when the structural, functional, and evolutionary evidence is considered together, it fits better with adaptation in laboratory cell culture, where immune and stomach constraints are missing, than with a long natural journey through animals. Acknowledgements I would like to thank Yuri Deigin, Daoyu Zhang, Steven Massey, Valentin Bruttel and Gilles Demaneuf for their insights and help while I prepared this paper.