(A) Schematic showing construction of recombinant SARS-CoV-2 harboring Venus and Cre in place of the ORF7a gene. (B) Schematic showing Venus and Cre expression. (C) Schematic of lineage tracing using recombinant viruses. Infected cells are Venus+ (green) at early times after infection, while surviving cells are tdTomato+ (red). (D) Infected VeroE6 cells (MOI = 0.01) were analyzed for GFP expression (top) and by bright-field microscopy (bottom) at 48 hours after infection (original magnification, ×20). (E) Mice were infected with the indicated viruses (K18-hACE2/Ai9 mice, 3,000 PFU rSARS-2-WH-V2C; Ai9 mice, 3,000 PFU rSARS-2-MA30-V2C) and monitored for survival (n = 5 mice per group). (F) Pathological changes in lungs of mice infected with rSARS-2-WH-V2C or rSARS-2-MA30-V2C at 4 dpi. Asterisks, hemorrhage; red arrowheads, edema; black arrowheads, perivascular infiltrates. Scale bar: 100 μm. (G) Sections from the indicated organs harvested from rSARS2-MA30-V2C–infected Ai9 mice were analyzed for Venus expression at 2 dpi. Venus+ cells were observed in the intestine (white arrows), nasal cavity, and lung. Scale bar: 50 μm. (H and I) Sections from the nasal cavities and lungs of rSARS2-MA30-V2C–infected Ai9 mice were analyzed for tdTomato expression at 20 (H) and 60 dpi (I). (J) Summary of numbers of tdTomato+ cells in the lungs of rSARS2-MA30-V2C–infected Ai9 mice. Each group contain 5 animals (n > 3 sections per animal were analyzed). (K) Rare tdTomato+ cells were found in the brains and hearts of rSARS2-WH-V2C–infected K18-hACE2/Ai9 mice at 20 dpi. Arrowheads indicate surviving cells. In D, F, and G–K images are representative of 2 independent experiments (5 mice/group, n = 4–10 slides per mouse). Higher magnification images of selected areas are shown in insets (G and H). Scale bars: 50 μm (G–K).