Fig. 6: SARS-CoV-2 detection in the brain.

Representative single-label IHC shows infrequent SARS-CoV-2 nucleocapsid (SARS-N) positivity in a cerebellar blood vessel of RM1 (a). Positivity was not detected in non-infected controls (AGM6 cerebellum b). SARS-CoV-2 spike (SARS-S) mRNA expression was assessed with in situ hybridization (RNAscope) but not seen in control animal tissue (RM6 cerebellum f-h). Rare positivity was seen in infection (AGM1 cerebellum c–e). RNAscope was performed 7 times on the brain regions investigated. A majority of brain tissue does not show any virus in infected animals which can be seen in the neighboring vessel (RM3 basal ganglia l–n) to a vessel with suggestive virus positivity (i–k). Endothelial cell infection is suggested by double fluorescent labeling of SARS-N with von Willebrand factor (vWF) (RM3 basal ganglia i–k). Merged images show colocalization of SARS-N (red; j) with vWF (green; i), indicated by white arrows. Blue color represents DAPI labeled cell nuclei. Immunohistochemical staining for SARS-N was performed 12 times on the brain regions investigated. SARS-CoV-2 RNA was detected in the different brain regions of infected animals via a CRISPR-based fluorescent method (o) where n = 3 repeats of independent samples. Dotted line indicates the cut-off value of positivity equal to 3.6 × 10⁶ photoluminescence (PL) intensity. Data are expressed as mean ± SEM. Source data are provided as a Source Data file. Abbreviations: BG basal ganglia, CER cerebellum, BS brainstem, FL frontal lobe, CSF cerebrospinal fluid, BC blank control, arb. units arbitrary units, AGM African green monkey, RM Rhesus macaque. Scale bars = 50 µm (a and b), 20 µm (c–h), and 10 µm (i–n). Fluorescence images are at 100×.