Vesicle-Cloaked Rotavirus Clusters are Environmentally Persistent and Resistant to Free Chlorine Disinfection

Recent discovery of vesicle-cloaked virus clusters (i.e., viral vesicles) has greatly challenged the central paradigm of viral transmission and infection as a single virion. To understand the environmental transmission of viral vesicles, we used an in vivo model to investigate their environmental persistence and engineering control by disinfection. Murine rotavirus vesicles maintained both their integrity and infectivity after incubation in filtered freshwater and wastewater for at least 7 days, with 24.5-27.5% of the vesicles still intact at 16 weeks after exposure to both waters. Free chlorine disinfection at a dosage of 13.3 mg min L^-1 did not decompose murine rotavirus vesicles, and it was much less effective in inactivating rotaviruses inside vesicles than free rotaviruses based on the quantification of rotavirus shedding in mouse stool and rotavirus replication in small intestines. Rotavirus vesicles may be more environmentally transmissible than free rotaviruses regardless of disinfection. Vesicle-mediated en bloc transmission could be responsible for vesicles' resistance to disinfection due to an increased multiplicity of infection and/or genetic recombination or reassortment to promote infection. Our work highlights the environmental, biological, and public health significance of viral vesicles, and the findings call for urgent action in advancing disinfection for pathogen control.