Measles virus exits human airway epithelia via infectious center sloughing

Measles virus (MeV) is the most contagious human virus, but we do not fully understand why. Unlike most respiratory viruses, MeV does not infect the airway epithelium immediately. MeV traverses the epithelium within immune cells that carry it to lymphatic organs where amplification occurs. Infected immune cells then synchronously deliver large amounts of virus to the airways. However, our understanding of MeV replication in airway epithelia is limited. To model it, we use well-differentiated primary cultures of human airway epithelial cells (HAE) from lung donors. In HAE, MeV spreads directly cell-to-cell forming infectious centers that grow for [~]3-5 days, are stable for a few days, and then disappear. Transepithelial electrical resistance remains intact during the entire course of HAE infection, thus we hypothesized that MeV infectious centers may slough off while preserving epithelial function. After documenting by confocal microscopy that infectious centers progressively detach from HAE, we recovered apical washes and separated cell-associated from cell-free virus by centrifugation. Virus titers were about 10 times higher in the cell-associated fraction than in the supernatant. In sloughed infectious centers, ciliary beating persisted and apoptotic markers were not readily detected, suggesting that they retain functional metabolism. Cell-associated MeV infected primary human monocyte-derived macrophages, modeling the first stage of infection in a new host. Single-cell RNA sequencing identified wound healing, cell growth, and cell differentiation as biological processes relevant for infectious center sloughing. 5-ethynyl-2-deoxyuridine (EdU) staining located proliferating cells underneath infectious centers. Thus, cells located below infectious centers divide and differentiate to repair the extruded infected epithelial patch. As an extension of these studies, we postulate that expulsion of infectious centers through coughing and sneezing could contribute to MeVs strikingly high reproductive number by allowing the virus to survive longer in the environment and by delivering a high infectious dose to the next host. AUTHOR SUMMARYMeasles virus (MeV) is a respiratory pathogen that infects millions worldwide each year. Although sometimes mischaracterized as an innocuous childhood disease, measles remains a leading cause of death for children under five. MeV is the most contagious human virus and requires vaccination rates above 90% to maintain herd immunity. Global decreases in vaccination rates over the past ten years contributed to recent, widespread MeV outbreaks. We uncover here a novel mechanism by which MeV exits the human airways that may explain why it is much more contagious than other viruses. We document that infected cells containing cell-associated virus slough en masse from the airway epithelial sheet. These expelled infectious centers are metabolically active and can transmit infection to primary human monocyte-derived macrophages more efficiently than cell-free virus particles. Thus, cell-associated MeV can transmit host-to-host, a new paradigm for efficient respiratory virus transmission.