Early innate immune response and evolution of a SARS-CoV-2 furin cleavage site inactive variant in bat cells

SARS-CoV-2 has caused the largest known coronavirus pandemic and is believed to have emerged from insectivorous bats. Little is known about the evolution of these viruses in their reservoir bat species. In this study, we investigated SARS-CoV-2-host interaction using human and bat cells. Bat cells mount a robust and early antiviral response but elicit a dampened pro-inflammatory response upon SARS-CoV-2 infection compared to human cells. Furthermore, an inactivating R685P mutation within the furin cleavage site (FCS) of the SARS-CoV-2 spike protein was naturally selected for in infected bat cells. Taken together, our data demonstrate that insectivorous bat cells have evolved a differential antiviral immune response against SARS-CoV-2 infection, likely to mitigate immunopathology that is observed in humans. Our study sheds light on the evolution of sarbecoviruses in bats and extends molecular evidence to data from field studies that have demonstrated that SARS-CoV-2-related viruses in wild-caught bats lack an intact FCS. Clinical isolate of ancestral SARS-CoV-2 (SARS-CoV-2/SB2) was used for infections. Cells at ~80% confluence were infected with SARS-CoV-2 at indicated multiplicity of infection (MOI). Control cells were treated with serum-free DMEM alone (mock infected). Infected and mock infected cells were incubated at 37 °C for 1 h with gentle rocking every 15 min. After 1 h, the virus inoculum was aspirated, cells were washed 1x with phosphate-buffered saline (PBS), and complete growth medium was added. All work with infectious SARS-CoV-2 was done in BSL3 laboratories at the Vaccine and Infectious Disease Organization (VIDO), University of Saskatchewan, Canada and McMaster University, Canada following approved protocols. Cells were infected with SARS-CoV-2 at an MOI of 0.1 for 12 and 24 h. Time-matched, sham infected cells served as controls. RNA was isolated from whole cell lysates using RNeasy Mini kit (Qiagen, 74106). Sequencing was conducted at the McMaster Genomics Facility, Farncombe Institute at McMaster University.