Cellular immunophenotyping in human and primate tissues during healthy conditions and Ebola and Nipah infections

We developed a 29-color spectral cytometry panel to enhance nonhuman primate (NHP) models for cross-reactive immunophenotyping. This panel is suitable for biosafety level 4 (BSL-4) viruses and can be used with both human and NHP samples in BSL-2 research settings. Tissues from humans, rhesus monkeys (RhMs), crab-eating macaques (CEMs), and green monkeys (GMs) were stained with a 29-color immunophenotyping panel requiring only 2 clone substitutions. Comparable staining was observed for all samples. Unbiased analysis showed acceptable overlap in T cell phenotypes across samples, with differences in human and NHP B cells and granulocytes. In CEMs, most circulating CD8+ T cells were from effector memory cells, with significantly higher levels than in humans, RhMs, and GMs. Analysis of samples from various anatomical sites revealed distinct location-specific phenotypes. In Nipah virus-exposed animals, splenocytes showed a substantial increase in IgM+ B cells and a reduction in effector memory CD8+ T cells compared with unexposed controls. Lymph nodes from Ebola virus-exposed animals showed a loss of CXCR3+CD8+ T cells versus unexposed controls. This panel may guide the development of additional multicolor panels in preclinical and clinical settings and may increase understanding of the pathogenesis of diseases caused by emerging and reemerging viruses.