Fig. 1. Experimental design and sample collection.

Seven adult Chinese-origin rhesus macaques (M0 to M6) were enrolled in the current study. At the outset of this experiment, six monkeys (M1 to M6) were challenged intratracheally with SARS-CoV-2 at 1 × 10⁶ TCID₅₀. After all the experimentally-infected monkeys had recovered from the primary infection, four infected monkeys (M3 to M6) were intratracheally rechallenged at 28 days post initial challenge (dpi) with the same dose of the SARS-CoV-2 strain, to ascertain the possibility of reinfection. In addition, an uninfected monkey (M0) was also treated with SARS-CoV-2 as the model control of the second challenge, and a previously infected monkey (M2) was untreated in the rechallenge experiment and was continuously monitored as the control animal. To compare the virus distribution and histopathological changes between the initially infected monkeys and the reinfected monkeys, two monkeys per group (M0 and M1 in the initial infection group, M3 and M5 in the reinfection group) were euthanized and necropsied at 5 dpi (M0), 7 dpi (M1) and 5 days post rechallenge (dpr) (M3 and M5), respectively. Body weight, body temperature, nasal/throat/anal swabs, hematological changes, immunocytes, and specific antibodies were measured over the short-term observation period. Two measurements of virus distribution and histopathology (H&E/IHC staining) were carried out at 5 dpi (M0), 7 dpi (M1), and 5 dpr (M3 and M5). Chest X-ray and neutralizing antibody titers against SARS-CoV-2 were examined at the indicated time points.