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Single-cell longitudinal analysis of SARS-CoV-2 infection in human airway epithelium

Neal G. RavindraiD, Mia Madel AlfajaroiD, Victor GasqueiD, Victoria Habet, Jin WeiiD, Renata B. FilleriD, Nicholas C. HustoniD, Han WaniD, Klara Szigeti-Buck, Bao WangiD, Guilin WangiD, Ruth R. MontgomeryiD, Stephanie C. EisenbarthiD, Adam WilliamsiD, Anna Marie PyleiD, Akiko IwasakiiD, Tamas L. Horvath, Ellen F. FoxmaniD, Richard W. PierceiD, David van DijkiD, Craig B. WileniD

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Abstract

SARS-CoV-2, the causative agent of COVID-19, has tragically burdened individuals and institutions around the world. There are currently no approved drugs or vaccines for the treatment or prevention of COVID-19. Enhanced understanding of SARS-CoV-2 infection and pathogenesis is critical for the development of therapeutics. To reveal insight into viral replication, cell tropism, and host-viral interactions of SARS-CoV-2 we performed single-cell RNA sequencing of experimentally infected human bronchial epithelial cells (HBECs) in air-liquid interface cultures over a time-course. This revealed novel polyadenylated viral transcripts and highlighted ciliated cells as a major target of infection, which we confirmed by electron microscopy. Over the course of infection, cell tropism of SARS-CoV-2 expands to other epithelial cell types including basal and club cells. Infection induces cell-intrinsic expression of type I and type III IFNs and IL6 but not IL1. This results in expression of interferon-stimulated genes in both infected and bystander cells. We observe similar gene expression changes from a COVID-19 patient ex vivo. In addition, we developed a new computational method termed CONditional DENSity Embedding (CONDENSE) to characterize and compare temporal gene dynamics in response to infection, which revealed genes relating to endothelin, angiogenesis, interferon, and inflammation-causing signaling pathways. In this study, we conducted an in-depth analysis of SARS-CoV-2 infection in HBECs and a COVID-19 patient and revealed genes, cell types, and cell state changes associated with infection.

Abstract summary

Claude Opus 5.5 · xhigh effort · 18 Oct 2026 · from the abstract · CC BY-NC 4.0

Time-course single-cell RNA sequencing of infected human bronchial epithelial cultures reports ciliated cells as a major target, with tropism later widening to basal and club cells.

The authors note that no approved drugs or vaccines for COVID-19 existed, making the cellular targets and host response of SARS-CoV-2 in the airway a prerequisite for therapeutic work. A time course in differentiated air-liquid interface cultures follows tropism and the interferon response together, with an ex vivo patient sample for comparison.

Key results

  1. ClaimedCiliated cells are reported as a major target of SARS-CoV-2 in bronchial epithelial cultures, with infection spreading over time to other epithelial types including basal and club cells.

    From single-cell RNA sequencing of experimentally infected air-liquid interface cultures sampled over a time course; ciliated-cell infection was checked by electron microscopy. No infected-cell fractions or per-type rates are given, and the authors also report polyadenylated viral transcripts they describe as previously unreported. · Abstract

  2. ClaimedInfection triggers cell-intrinsic type I and type III interferon and IL6 expression but not IL1, and interferon-stimulated genes appear in bystander cells as well as infected ones.

    Stated as an observation from the same cultures, with comparable expression changes reported in an ex vivo sample from one COVID-19 patient; no fold changes, cell counts or statistics are reported. · Abstract

  3. ClaimedA new method, CONditional DENSity Embedding (CONDENSE), compares temporal gene expression dynamics after infection and points to endothelin, angiogenesis, interferon and inflammatory signalling pathways.

    Described as developed for this study; the abstract gives no algorithmic detail, benchmark against existing trajectory or temporal-clustering methods, or enrichment statistics. · Abstract

Methods, data and reproducibility

Human bronchial epithelial cells grown at an air-liquid interface were experimentally infected with SARS-CoV-2 and profiled by single-cell RNA sequencing across a time course, with electron microscopy of infected cells, comparison with an ex vivo sample from a COVID-19 patient, and a purpose-built embedding method (CONDENSE) for temporal gene dynamics.

  • SARS-CoV-2-infected human bronchial epithelial cells (HBECs) in air-liquid interface culture, single-cell RNA sequencing over a time course · human; SARS-CoV-2
  • ex vivo sample from a COVID-19 patient used for comparison of expression changes · human · n = one patient
  • electron microscopy of infected airway cultures · human; SARS-CoV-2
Methods & tools
single-cell RNA sequencing, CONditional DENSity Embedding (CONDENSE), electron microscopy, air-liquid interface culture
Reproducibility gaps
No accession for the sequencing data and no repository for the CONDENSE implementation appear in the abstract.
Limitations (2)
  • The in vivo comparison rests on a single ex vivo COVID-19 patient sample, so agreement between culture and patient is anecdotal at this level of reporting. · noted in this summary
  • No quantities are reported anywhere in the available text: no infected-cell fractions, differential expression statistics or evaluation of CONDENSE against existing temporal-analysis methods. · noted in this summary

Read this if you study airway infection or interferon responses at single-cell resolution and want a time-resolved account of SARS-CoV-2 tropism in differentiated cultures.

Citations
74

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