Uncovering strain- and age- dependent differences in innate immune response to SARS-CoV-2 infection in nasal epithelia using combined short and long-read scRNA-seq

Assessing the impact of SARS-CoV-2 variants on host immune systems is crucial with the continuous arrival of novel variants. However, there is a lack of reported studies utilizing single-cell RNA-sequencing (scRNA-seq) to elucidate the age-dependent host-viral interactions with different SARS-CoV-2 strains. Therefore, we employed Full-Length Transcriptome Sequencing (FLT-Seq) combining Illumina and Oxford Nanopore Technologies (ONT) with 10X 3-prime single cell sequencing to investigate host transcriptomic changes during wild-type (WT) and Alpha-strain SARS-CoV-2 infections within air-liquid-interface (ALI)-cultured human nasal epithelial cells from adults and adolescents. The results revealed increased innate immune responses in cells with lower viral loads which were lacking in cells with higher viral load. Alpha-infections showed heightened expression of genes related to interferon (IFN)-responses and protein-folding compared with WT, implying the increased immune response and endoplasmic reticulum stress with the variant of a higher transmission rate. ACE2 expression was elevated in infected populations of secretory and goblet cells with high IFN-stimulation and a subpopulation of ciliated cells but was lacking in bystander cells and other infected-cell types, despite detectable IFN-stimulation and regardless of strain. We used long-read sequencing to identify differential transcript usage (DTU) of IFN-response and translational genes, including IFI27, RPL4 and RPL15 (padj < 0.05) in infected cells compared with control, and consistent DTU of RPL15 and MX1 between Alpha and WT strain-infected cells in various cell-types. Together, these results reveal the dynamic transcriptional/translational/post-translational activity upon SARS-CoV-2 infection, which appeared to be age and strain-dependent. Overall, this study highlights the complexity of cell-type, age and viral-strain-dependent host epithelial responses to SARS-CoV-2. ### Competing Interest Statement LC and MP have received travel funding from Oxford Nanopore Technologies. LC owns stock in Oxford Nanopore Technologies