Comparison of SARS-CoV-2 variants of concern in primary human nasal cultures demonstrates Delta as most cytopathic and Omicron as fastest replicating.

The SARS-CoV-2 pandemic was marked with emerging viral variants, some of which were designated as variants of concern (VOCs) due to selection and rapid circulation in the human population. Here, we elucidate functional features of each VOC linked to variations in replication rate. Patient-derived primary nasal cultures grown at air-liquid interface were used to model upper respiratory infection and compared to cell lines derived from human lung epithelia. All VOCs replicated to higher titers than the ancestral virus, suggesting a selection for replication efficiency. In primary nasal cultures, Omicron replicated to the highest titers at early time points, followed by Delta, paralleling comparative studies of population sampling. All SARS-CoV-2 viruses entered the cell primarily via a transmembrane serine protease 2 (TMPRSS2)-dependent pathway, and Omicron was more likely to use an endosomal route of entry. All VOCs activated and overcame dsRNA-induced cellular responses, including interferon (IFN) signaling, oligoadenylate ribonuclease L degradation, and protein kinase R activation. Among the VOCs, Omicron infection induced expression of the most IFN and IFN-stimulated genes. Infections in nasal cultures resulted in cellular damage, including a compromise of cell barrier integrity and loss of nasal cilia and ciliary beating function, especially during Delta infection. Overall, Omicron was optimized for replication in the upper respiratory tract and least favorable in the lower respiratory cell line, and Delta was the most cytopathic for both upper and lower respiratory cells. Our findings highlight the functional differences among VOCs at the cellular level and imply distinct mechanisms of pathogenesis in infected individuals. IMPORTANCE: Comparative analysis of infections by SARS-CoV-2 ancestral virus and variants of concern, including Alpha, Beta, Delta, and Omicron, indicated that variants were selected for efficiency in replication. In infections of patient-derived primary nasal cultures grown at air-liquid interface to model upper respiratory infection, Omicron reached the highest titers at early time points, a finding that was confirmed by parallel population sampling studies. While all infections overcame dsRNA-mediated host responses, infections with Omicron induced the strongest interferon and interferon-stimulated gene response. In both primary nasal cultures and lower respiratory cell line, infections by Delta were most damaging to the cells as indicated by syncytia formation, loss of cell barrier integrity, and nasal ciliary function.