Extended Data Fig. 3: Accumulation of viral antigens in the brain parallels the development of type I interferon response and downregulation of microglial P2Y12R.

a. Viral mRNA levels were measured by qPCR and presented as log arbitrary units. One-way ANOVA with Tukey’s post-hoc test (all groups compared): ^*p<0.05, ^**p<0.01, ^***p<0.001, data from n=11 COVID-19 cases. b. Virus load shows a positive correlation with IFN-α mRNA levels in the gyrus rectus (individual samples where SARS-CoV-2 mRNA levels were detectable were included). Pearson correlation with P values and Pearson r correlation coefficient displayed, n=7. c. IFN-β and IFN-α mRNA levels in tissue homogenates as measured by qPCR. Kruskal-Wallis test with Dunn's multiple comparison (all groups compared): ^*p<0.05, ^**p<0.01, ^***p<0.001, ^****p<0.0001; Kruskal-Wallis test with Dunn's multiple comparison (gyrus rectus vs brain areas): ^#p<0.05, ^##p<0.01, ^###p<0.001, ^####p<0.0001, n=11 COVID-19 cases. d. SARS-CoV-2 nucleocapsid and S1 protein labelings in lung epithelial cells. Lack of primary antibody eliminates specific immnopositivity for viral antigens on subsequent lung tissue sections. e. Immunodetection of SARS-CoV-2 nucleocapsid (arrowheads) in paraffin embedded bulbus olfactorius with cresyl violet counterstain (left panels). A subset of microglia accumulate viral antigens as visualized by immunofluorescence at infected areas of the bulbus (right panels). f. (i) Cranial nerve IX presents SARS-CoV-2 nucleocapsid immunopositive profiles (arrowheads). (ii) Viral antigens also appear in the vicinity of neurons and other cells outside blood vessels in the dorsal medulla. g. Loss of P2Y12R from vessel-associated microglia and microglial processes (arrowhead) with some P2Y12R retained on microglia away from the vessel (arrow) is shown in the gyrus rectus in a COVID-19 case. h. Perivascular CD15 positive immune cell (green) containing SARS-CoV-2 nucleocapsid (arrowheads), with scattered nucleocapsid staining seen among GFAP-positive astrocyte endfeet. i. Perivascular CD45-positive leukocytes (white, arrowhead) without intracellular viral antigens associated with microglial processes (Iba1, yellow) in the vicinity of SARS-CoV-2–immunopositive cells (magenta, arrow). v- lumen of blood vessel. j. Perivascular CD45-positive immune cells (arrowheads) containing SARS-CoV-2 nucleocapsid are contacted and internalized by Iba-positive microglia (arrows) in the gyrus rectus. Scale bars: d: 100 µm, 10 µm; e: 10 µm; f: 100 µm, 5 µm; g-h: 10 µm; i: 5 µm a, c, Data are presented as mean ± SD.