scEbola

Single-cell profiling of Ebola virus infection, in vivo and ex vivo

Schematic of experimental design

Overview

This was a large collaboration using the Seq-Well technology from Alex Shalek’s lab at MIT and CyTOF from Garry Nolan’s lab at Stanford to profile circulating immune cells during Ebola virus infection in Rhesus Macaques. Experiments were conducted in a biosafetey level 4 facility at the Integrated Research Facility, the NIH’s BSL-4 research campus in Frederick, Maryland. We quantified both host and viral gene expression within single cells which allowed us to characterize viral tropism, identify expression changes associated with viral replication, and to describe the broad changes in the immune system that occur in this uniformly lethal model of disease.

Special thanks to Aaron Lin, my co-first author and experimental master-mind for this project, as well as to the many other authors who made this project a reality including James Logue, Travis Hughes, Alex Shalek, Garry Nolan, Lisa Hensley, Pardis Sabeti, Richard Bennett, and David McIlwain to name just a few.

Abstract

Ebola virus (EBOV) causes epidemics with high mortality yet remains understudied due to the challenge of experimentation in high-containment and outbreak settings. Here, we used single-cell transcriptomics and CyTOF-based single-cell protein quantification to characterize peripheral immune cells during EBOV infection in rhesus monkeys. We obtained 100,000 transcriptomes and 15,000,000 protein profiles, finding that immature, proliferative monocyte-lineage cells with reduced antigen-presentation capacity replace conventional monocyte subsets, while lymphocytes upregulate apoptosis genes and decline in abundance. By quantifying intracellular viral RNA, we identify molecular determinants of tropism among circulating immune cells and examine temporal dynamics in viral and host gene expression. Within infected cells, EBOV downregulates STAT1 mRNA and interferon signaling, and it upregulates putative pro-viral genes (e.g., DYNLL1 and HSPA5), nominating pathways the virus manipulates for its replication. This study sheds light on EBOV tropism, replication dynamics, and elicited immune response and provides a framework for characterizing host-virus interactions under maximum containment.

Highlights:

  • Interferon response is suppressed in infected cells but activated in bystander cells
  • EBOV represses antiviral genes and upregulates pro-viral genes in infected cells
  • Proliferative CD14– CD16– monocyte precursors expand in circulation during EVD
  • Identification of expression markers of EBOV tropism for circulating cells in vivo